A quantifiler™ trio-based HRM screening assay for the accurate prediction of single source versus mixed biological samples.

At present, the forensic DNA workflow is not capable of providing information about the contributor status (single source vs. multiple contributors) of evidentiary samples prior to end-point analysis. This exacerbates the challenges inherent to mixtures and low-template DNA samples. If additional sample information could be provided earlier in the workflow, protocols could be implemented to mitigate these challenges. An integrated Quantiplex®- high resolution melt (HRM) assay was shown to be effective in distinguishing between single source and mixture DNA samples; however, integration of the HRM assay into a more commonly used chemistry would be beneficial to the practitioner community. Thus, the assay was redesigned as an integrated Quantifiler™ Trio-HRM assay, which included the identification of a new DNA-binding dye, an increased reaction volume, and the establishment of new data analysis and standard curve metrics for all targets. This redesigned assay produced quantification values and qualitative values that were comparable to those produced when the same samples were tested using the standard Quantifiler™ Trio chemistry and settings. Further, STR profiles generated with quantification values produced from the integrated Quantifiler™ Trio-HRM assay and standard Quantifiler™ Trio chemistry were complete and fully concordant. Most importantly, the integrated Quantifiler™ Trio-HRM assay was able to accurately predict whether a sample was single source or a mixture 79.2% of the time, demonstrating the potential of this approach. With the incorporation of an expanded training set for prediction modeling, and completion of critical developmental validation studies, this assay could prove useful to the forensic DNA practitioner community.

Replication Protein A1 is essential for DNA damage repair during mammalian oogenesis.

Persistence of unrepaired DNA damage in oocytes is detrimental and may cause genetic aberrations, miscarriage, and infertility. RPA, an ssDNA-binding complex, is essential for various DNA-related processes. Here we report that RPA plays a novel role in DNA damage repair during postnatal oocyte development after meiotic recombination. To investigate the role of RPA during oogenesis, we inactivated RPA1 (replication protein A1), the largest subunit of the heterotrimeric RPA complex, specifically in oocytes using two germline-specific Cre drivers (Ddx4-Cre and Zp3-Cre). We find that depletion of RPA1 leads to the disassembly of the RPA complex, as evidenced by the absence of RPA2 and RPA3 in RPA1-deficient oocytes. Strikingly, severe DNA damage occurs in RPA1-deficient GV-stage oocytes. Loss of RPA in oocytes triggered the canonical DNA damage response mechanisms and pathways, such as activation of ATM, ATR, DNA-PK, and p53. In addition, the RPA deficiency causes chromosome misalignment at metaphase I and metaphase II stages of oocytes, which is consistent with altered transcript levels of genes involved in cytoskeleton organization in RPA1-deficient oocytes. Absence of the RPA complex in oocytes severely impairs folliculogenesis and leads to a significant reduction in oocyte number and female infertility. Our results demonstrate that RPA plays an unexpected role in DNA damage repair during mammalian folliculogenesis.

CYP eicosanoid pathway mediates colon cancer-promoting effects of dietary linoleic acid.

Human and animal studies support that consuming a high level of linoleic acid (LA, 18:2ω-6), an essential fatty acid and key component of the human diet, increases the risk of colon cancer. However, results from human studies have been inconsistent, making it challenging to establish dietary recommendations for optimal LA intake. Given the importance of LA in the human diet, it is crucial to better understand the molecular mechanisms underlying its potential colon cancer-promoting effects. Using LC-MS/MS-based targeted lipidomics, we find that the cytochrome P450 (CYP) monooxygenase pathway is a major pathway for LA metabolism in vivo. Furthermore, CYP monooxygenase is required for the colon cancer-promoting effects of LA, since the LA-rich diet fails to exacerbate colon cancer in CYP monooxygenase-deficient mice. Finally, CYP monooxygenase mediates the pro-cancer effects of LA by converting LA to epoxy octadecenoic acids (EpOMEs), which have potent effects on promoting colon tumorigenesis via gut microbiota-dependent mechanisms. Overall, these results support that CYP monooxygenase-mediated conversion of LA to EpOMEs plays a crucial role in the health effects of LA, establishing a unique mechanistic link between dietary fatty acid intake and cancer risk. These results could help in developing more effective dietary guidelines for optimal LA intake and identifying subpopulations that may be especially vulnerable to LA's negative effects.

Early embryonic lethality of mice lacking POLD2.

As a highly conserved DNA polymerase (Pol), Pol δ plays crucial roles in chromosomal DNA synthesis and various DNA repair pathways. However, the function of POLD2, the second small subunit of DNA Pol δ (p50 subunit), has not been characterized in vivo during mammalian development. Here, we report for the first time, the essential role of subunit POLD2 during early murine embryogenesis. Although Pold2 mutant mouse embryos exhibit normal morphology at E3.5 blastocyst stage, they cannot be recovered at gastrulation stages. Outgrowth assays reveal that mutant blastocysts cannot hatch from the zona pellucida, indicating impaired blastocyst function. Notably, these phenotypes can be recapitulated by small interfering RNA (siRNA)-mediated knockdown, which also exhibit slowed cellular proliferation together with skewed primitive endoderm and epiblast allocation during the second cell lineage specification. In summary, our study demonstrates that POLD2 is essential for the earliest steps of mammalian development, and the retarded proliferation and embryogenesis may also alter the following cell lineage specifications in the mouse blastocyst embryos.

A brief look at diagnosing neurocognitive disorders in a VA primary care setting: Understanding the practices of our physician partners and the future of integration.

OBJECTIVE: The population aged 65 and over is growing rapidly and with it are increased demands for integrative care and management of cognitive health issues. Such care models often do not include neuropsychologists or other psychology specialties. METHODS: In this study, 72 medical charts of VA patients diagnosed with neurocognitive disorders were reviewed using best practice guidelines for diagnosing and managing neurocognitive disorders, adapted from Downs et al. (2006). RESULTS: Results indicated that physicians typically used clinical judgment through review of medications, blood work within the past year, and a history of symptoms to make diagnostic conclusions. Cognitive assessment and consideration of other reversible causes of cognitive decline (e.g., depression) were less commonly considered. CONCLUSIONS: The results are discussed in terms of potential implications for neuropsychologists and the integration of neuropsychology and primary care.

Examining the factor structure, reliability, and validity of the main outcome measure used in mentalization-based therapy skills training.

BACKGROUND AND AIMS: Multiple national guidelines stress the importance for clinicians to possess good therapeutic skills for working with patients with significant relational difficulties (who may receive a diagnosis of personality disorder). Training clinicians in mentalization-based treatment skills (MBT-S) is one approach to address this. The main outcome measure used in MBT-S studies is the Knowledge and Application of MBT Questionnaire (KAMQ). However, an absence of research into the properties and validity of the KAMQ has limited the methodological quality of MBT-S evaluations so far. The aim of this study was therefore to investigate the factor structure, internal consistency, reliability, and validity of the KAMQ. METHODS: Using an existing multiprofessional sample of 217 clinicians from 2014 to 2016, we undertook exploratory factor analysis to determine the factor structure and internal consistency of the KAMQ. Convergent validity of the measure with the Attitudes to Personality Disorder Questionnaire (APDQ) was assessed in a subset of this dataset where both questionnaires had been administered (n = 92). Additionally, by recruiting a new sample of 70 clinicians, we assessed the measure's test-retest reliability. RESULTS: Factor analysis found three factors underlying 17 of the 20 KAMQ items, relating to therapeutic skills in mentalizing, beliefs about applying MBT in practice, and specific MBT knowledge. The KAMQ was revised following the factor analysis to form the KAMQ-2 with 17 items. Internal consistency (α = .85, 95% confidence interval [CI] = 0.80-0.89) and test-retest reliability (ICC = 0.84, 95% CI = 0.73-0.91) were good. In correlation analyses, the KAMQ-2 showed convergent validity with the main factor from the APDQ (n = 48; r s = 0.39, P < .01). CONCLUSION: The KAMQ-2 provides a short, reliable self-report instrument which probes clinicians' knowledge about mentalizing skills, and beliefs about using these. There was preliminary evidence for validity. The properties of the KAMQ-2 mean that more robust evaluation and development of MBT-S is now possible.

Associations of Future Cognitive Decline with Sexual Satisfaction among Married Older Adults.

Objectives: This study sought to explore changes in longitudinal cognitive status in relation to baseline measures of intimacy and sexuality in cognitively intact, married older adults.Methods: Baseline intimacy and sexuality survey data from 155, cognitively intact, married, older adults were collected using a novel survey instrument that explored the domains of: 1) romance with one's partner, 2) sexual satisfaction, 3) beliefs about sexuality, and 4) social support and emotional intimacy. These data were analyzed in relation to change in cognitive status over a 10-year follow-up period using binary logistic regression modeling. Exploratory factor analysis was used to assess the shared variance of survey items attributable to intimacy and sexuality without specification of an a priori hypothesis regarding the association of intimacy and sexuality with future change in cognitive status.Results: Over the 10-year study period, 33.5% (n = 52) of individuals developed cognitive impairment. Participants with greater sexual satisfaction scores at baseline were statistically less likely to convert from cognitively intact to mild cognitive impairment or dementia in the future (p = .01). The domains of romance with one's partner, beliefs about sexuality, and social support/emotional intimacy were not predictive of future longitudinal changes in cognitive status.Conclusions: Sexual satisfaction is associated with longitudinal cognitive outcomes in cognitively intact, married, older adults.Clinical implications: Clinicians should routinely assess for sexual satisfaction among older adults and refer to appropriate providers, such as couples or sex therapists, when appropriate.

One Health Genomic Study of Human and Animal Klebsiella pneumoniae Isolated at Diagnostic Laboratories on a Small Caribbean Island.

Klebsiella pneumoniae causes a variety of infections in both humans and animals. In this study, we characterised the genomes of human and animal isolates from two diagnostic laboratories on St. Kitts, a small Caribbean island inhabited by a large population of vervet monkeys. In view of the increased chances of direct or indirect contact with humans and other animal species, we used the One Health approach to assess transmission of K. pneumoniae across host species by sequencing 82 presumptive K. pneumoniae clinical isolates from humans (n = 51), vervets (n = 21), horses (n = 5), dogs (n = 4) and a cat (n = 1). Whole genome sequencing (WGS) was carried out using Illumina technology. De novo assembly was performed in CLC Genomics Workbench v.11.0. Single nucleotide polymorphisms were detected using NASP followed by phylogenetic analysis using IQ-TREE. Virulence and antimicrobial resistance gene contents were analysed using the Kleborate and CGE pipelines. WGS-based analysis showed that 72 isolates were K. pneumoniae sensu stricto and five K. quasipneumoniae and five K. variicola. K. pneumoniae isolates belonged to 35 sequence types (ST), three of which were occasionally shared between humans and animals: ST23, ST37 and ST307. The ST23 strains from vervets formed a separate cluster amongst publicly available sequenced ST23 strains, indicating the presence of a specific vervet sublineage. Animal strains harbored fewer resistance genes and displayed distinct virulence traits that appeared to be host-specific in vervet isolates. Our results show that K. pneumoniae infections on this Caribbean island are usually caused by host-specific lineages.

Pharmacy-facilitated medication history program at a community teaching hospital: A pre-post study in an emergency department.

PURPOSE: A study was conducted to compare the accuracy of medication histories compiled by pharmacy technicians with histories obtained through the usual multidisciplinary process. METHODS: A retrospective cohort study was conducted at a community teaching hospital from January 2017 through February 2018. Inclusion criteria included patient age of at least 18 years, use of 1 or more medications at the time of admission, and hospital admission through the emergency department. Each electronically documented medication history was assessed for accuracy. The objective was to compare the accuracy of pharmacy technician-collected medication histories to those obtained through the usual multidisciplinary process. RESULTS: Of 215 patients screened, 183 were included in the study: 91 patients whose medication histories were obtained through the usual multidisciplinary process and 92 whose medication histories were collected by pharmacy technicians. Overall, documentation for 1,773 medications listed in medication histories was reviewed. The primary outcome of medication history accuracy occurred 38% of the time with the usual multidisciplinary process and 70% of the time with pharmacy technician collection of medication histories (P < 0.001). CONCLUSION: The study showed that the accuracy of medication histories was improved when histories were obtained by pharmacy technicians instead of via the usual multidisciplinary process.

High Prevalence of USA300 Among Clinical Isolates of Methicillin-Resistant Staphylococcus aureus on St. Kitts and Nevis, West Indies.

Limited information is available on antimicrobial susceptibility and clonal distribution of Staphylococcus aureus in the Caribbean region. The aims of this study were to determine the prevalence of antimicrobial resistance among S. aureus isolates and to reveal the frequency and population structure of methicillin-resistant S. aureus (MRSA) in St. Kitts and Nevis, a small island country in the West Indies. A total of 152 S. aureus isolates were collected from consecutive samples submitted to the clinical microbiology laboratory of the main referral hospital from March 2017 to January 2018. Samples came from all units in the hospital and a small number came from external submissions, and comprised a total of 119 clinical specimens and 33 nasal swabs collected from staff and patients. All S. aureus isolates were confirmed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Minimal Inhibitory Concentrations (MICs) of clinically relevant antimicrobials were determined by broth microdilution, and diversity of MRSA isolates was assessed by whole genome sequencing (WGS) analysis. MRSA accounted for 45% (69/152) of the isolates. The highest rates of resistance to non-ß-lactam agents were observed for erythromycin (55%), moxifloxacin (41%), and levofloxacin (40%), whereas resistance to the other drugs tested was ≤6%. All isolates were susceptible to ceftaroline, linezolid, teicoplanin, telavancin, and vancomycin. WGS-based multilocus sequence typing (MLST) showed that approximately 88% of the MRSA isolates belonged to ST8. Phylogenetic analysis on 801 single-nucleotide polymorphisms (SNPs) identified among the MRSA ST8 isolates indicates a large degree of genetic diversity. However, all ST8 strains clustered within the distinct clade that defines the USA300 North American Epidemic lineage (Panton-Valentine Leukocidin (PVL) +, arginine catabolic mobile element (ACME) +, Staphylococcal cassettes chromosome mec IVa (SCCmec IVa)). Our data show high levels of methicillin, macrolide, and fluoroquinolone resistance among S. aureus on St. Kitts and Nevis. The USA300 North American epidemic lineage is responsible for the vast majority of MRSA infections on this Caribbean island.

Forgotten, silent, and overruled: Hearing the voice of the nursing home patient.

As the population ages, more Americans are moving into nursing homes/long-term care facilities. Per Accreditation Council for Graduate Medical Education 2017 guidelines, family medicine residents are required to gain experience and competence working in long-term care facilities; however, this unique environment poses several challenges for residents to hear the wishes of their patients over the demands of the patient's medical care team and family members. Also, many patients in long-term care facilities have sensory impairments (e.g., poor eyesight, deafness) and/or cognitive deficits (e.g., dementia). One solution for hearing the patient's voice over the demands of medical professionals and family members is to train physicians on how to overcome communication barriers with their patients. This article will discuss solution-focused approaches to reducing the barriers of sensory and cognitive impairments through the use of adaptive communication behaviors and adaptive equipment. Ways to improve communication between physicians, nursing-home staff, and patients' family in order to improve the care patients receive in long-term care facilities will also be addressed. These recommendations are designed to assist with reducing physician frustration, increasing each patient's input in medical decision-making, and improving communication across the patient care team.

Nontyphoidal Salmonella enterica Nonsusceptible to Both Levofloxacin and Ceftriaxone in Nebraska, United States 2014-2015.

Nontyphoidal Salmonella enterica is a common cause of illness in humans ranging from gastroenteritis to invasive disease. National surveillance programs continually monitor trends in antimicrobial resistance patterns and mechanisms of resistance to identify emerging public health threats. Our study shows the emergence of nonsusceptibility to both levofloxacin and ceftriaxone, a concerning phenotype that threatens first-line antibiotic therapy, in Salmonella isolates recovered between 2014 and 2015. From 2010 to 2013 the rate of resistance increased from 0.0% (0/1181) to 1.5% (9/593) in 2014 and 2015. The isolates with this phenotype were found to be from multiple serotypes, including Typhimurium, Newport, and Enteritidis. Resistance to ceftriaxone was attributed to the presence of either an AmpC or extended-spectrum ß-lactamase, and resistance to fluoroquinolones was attributable to the presence of mutations in the quinolone resistance-determining region or the presence of plasmid-mediated quinolone resistance genes. As this resistance pattern was seen in a variety of Salmonella serotypes harboring varied resistance mechanisms, it indicates a worrying trend in the spread of isolates resistant to both first-line treatment options.

Cochlear implantation for single-sided deafness and tinnitus suppression.

OBJECTIVE: To quantify the potential effectiveness of cochlear implantation for tinnitus suppression in patients with single-sided deafness using the Tinnitus Handicap Inventory. METHODS: The study included 12 patients with unilateral tinnitus who were undergoing cochlear implantation for single-sided deafness. The Tinnitus Handicap Inventory was administered at the patient's cochlear implant candidacy evaluation appointment prior to implantation and every cochlear implant follow-up appointment, except activation, following implantation. Patient demographics and speech recognition scores were also retrospectively recorded using the electronic medical record. RESULTS: A significant reduction was found when comparing Tinnitus Handicap Inventory score preoperatively (61.2±27.5) to the Tinnitus Handicap Inventory score after three months of cochlear implant use (24.6±28.2, p=0.004) and the Tinnitus Handicap Inventory score beyond 6months of CI use (13.3±18.9, p=0.008). Further, 45% of patients reported total tinnitus suppression. Mean CNC word recognition score improved from 2.9% (SD 9.4) pre-operatively to 40.8% (SD 31.7) by 6months post-activation, which was significantly improved from pre-operative scores (p=0.008). CONCLUSION: The present data is in agreement with previously published studies that have shown an improvement in tinnitus following cochlear implantation for the large majority of patients with single-sided deafness.

Tip Fold-over in Cochlear Implantation: Case Series.

OBJECTIVE: To describe the incidence, clinical presentation, and performance of cochlear implant (CI) recipients with tip fold-over. STUDY DESIGN: Retrospective case series. SETTING: Tertiary referral center. PATIENTS: CI recipients who underwent postoperative computed tomography (CT) scanning. INTERVENTION(S): Tip fold-over was identified tomographically using previously validated software that identifies the electrode array. Electrophysiologic testing including spread of excitation or electric field imaging (EFI) was measured on those with fold-over. MAIN OUTCOME MEASURE(S): Location of the fold-over; audiological performance pre and postselective deactivation of fold-over electrodes. RESULTS: Three hundred three ears of 235 CI recipients had postoperative CTs available for review. Six (1.98%) had tip fold-over with 5/6 right-sided ears. Tip fold-over occurred predominantly at 270 degrees and was associated with precurved electrodes (5/6). Patients did not report audiological complaints during initial activation. In one patient, the electrode array remained within the scala tympani with preserved residual hearing despite the fold-over. Spread of excitation supported tip fold-over, but the predictive value was not clear. EFI predicted location of the fold-over with clear predictive value in one patient. At an average follow-up of 11 months, three subjects underwent deactivation of the overlapping electrodes with two of them showing marked audiological improvement. CONCLUSION: In a large academic center with experienced surgeons, tip fold-over occurred at a rate of 1.98% but was not immediately identifiable clinically. CT imaging definitively showed tip fold-over. Deactivating involved electrodes may improve performance possibly avoiding revision surgery. EFI may be highly predictive of tip fold-over and can be run intraoperatively, potentially obviating the need for intraop fluoroscopy.

A framework for assessing outcomes from newborn screening: on the road to measuring its promise.

UNLABELLED: Newborn screening (NBS) is intended to identify congenital conditions prior to the onset of symptoms in order to provide early intervention that leads to improved outcomes. NBS is a public health success, providing reduction in mortality and improved developmental outcomes for screened conditions. However, it is less clear to what extent newborn screening achieves the long-term goals relating to improved health, growth, development and function. We propose a framework for assessing outcomes for the health and well-being of children identified through NBS programs. The framework proposed here, and this manuscript, were approved for publication by the Secretary of Health and Human Services' Advisory Committee on Heritable Disorders in Newborns and Children (ACHDNC). This framework can be applied to each screened condition within the Recommended Uniform Screening Panel (RUSP), recognizing that the data elements and measures will vary by condition. As an example, we applied the framework to sickle cell disease and phenylketonuria (PKU), two diverse conditions with different outcome measures and potential sources of data. Widespread and consistent application of this framework across state NBS and child health systems is envisioned as useful to standardize approaches to assessment of outcomes and for continuous improvement of the NBS and child health systems. SIGNIFICANCE: Successful interventions for newborn screening conditions have been a driving force for public health newborn screening for over fifty years. Organizing interventions and outcome measures into a standard framework to systematically assess outcomes has not yet come into practice. This paper presents a customizable outcomes framework for organizing measures for newborn screening condition-specific health outcomes, and an approach to identifying sources and challenges to populating those measures.

Strengthening partnerships between Black Churches and HIV service providers in the United States.

Across the United States, Black Churches play a significant role among the Black community and are increasingly being used to deliver Human Immunodeficiency Virus (HIV) prevention services. This study sought to investigate HIV service providers' strategies for strengthening partnerships with churches to deliver HIV prevention services. Using a community-based participatory research approach, an HIV coalition and an academic institution formed a partnership to address the study aim. Individual interviews (n = 16) were conducted with providers from medical institutions and HIV social support agencies. A thematic analysis focusing on recommendations for addressing the challenges and benefits of partnership with churches for HIV services was conducted. Participants' interest in and intention to work with churches, as well as their comfort level discussing sexual health-related topics with religious congregations, was high. Four themes emerged to highlight the different perspectives of service providers' recommendations for addressing challenges and strengthening partnerships with churches to deliver HIV services including: (1) carefully selecting churches and HIV services to provide, (2) gaining "buy-in" and support of church leadership, (3) taking advantage of conflict with church doctrine, and (4) choosing appropriate delivery strategies. Study findings demonstrate that although challenges exist, heath service providers in this region of the United States may be interested in addressing HIV among faith communities. Study findings also provide concrete solutions to previously documented barriers to HIV prevention in Black Churches. Such information will benefit researchers and practitioners seeking to expand effective HIV prevention efforts with Black Churches in communities who bear a disproportionate burden of HIV infections.