Wastewater based surveillance can be used to reduce clinical testing intensity on a university campus.

Clinical testing has been a vital part of the response to and suppression of the COVID-19 pandemic; however, testing imposes significant burdens on a population. College students had to contend with clinical testing while simultaneously dealing with health risks and the academic pressures brought on by quarantines, changes to virtual platforms, and other disruptions to daily life. The objective of this study was to analyze whether wastewater surveillance can be used to decrease the intensity of clinical testing while maintaining reliable measurements of diseases incidence on campus. Twelve months of human health and wastewater surveillance data for eight residential buildings on a university campus were analyzed to establish how SARS-CoV-2 levels in the wastewater can be used to minimize clinical testing burden on students. Wastewater SARS-CoV-2 levels were used to create multiple scenarios, each with differing levels of testing intensity, which were compared to the actual testing volumes implemented by the university. We found that scenarios in which testing intensity fluctuations matched rise and falls in SARS-CoV-2 wastewater levels had stronger correlations between SARS-CoV-2 levels and recorded clinical positives. In addition to stronger correlations, most scenarios resulted in overall fewer weekly clinical tests performed. We suggest the use of wastewater surveillance to guide COVID-19 testing as it can significantly increase the efficacy of COVID-19 surveillance while reducing the burden placed on college students during a pandemic. Future efforts should be made to integrate wastewater surveillance into clinical testing strategies implemented on college campuses.

IL-21-IgFc immunotherapy alters transcriptional landscape of lymph node cells leading to enhanced flu vaccine response in aging and SIV infection.

Aging people living with HIV (PWH) frequently manifest impaired antibody (Ab) responses to seasonal flu vaccination which has been attributed to ongoing inflammation and immune activation. We have recently reported a similar scenario in old simian immunodeficiency virus (SIV) infected rhesus macaques (RM) with controlled viremia and have been able to compensate for this deficiency by immunotherapy with interleukin (IL)-21-IgFc. To understand the underlying mechanisms of IL-21-induced immunomodulation leading to enhanced flu vaccine response in aging and SIV, we have investigated draining lymph node (LN) cells of IL-21-treated and -untreated animals at postvaccination. We observed IL-21-induced proliferation of flu-specific LN memory CD4 T cells, expansion of B cells expressing IL-21 receptor (IL-21R), and modest expansion of T follicular helper cells (Tfh) co-expressing T-cell immunoreceptor with Ig and ITIM domains (TIGIT) and DNAX accessory molecule (DNAM-1). Transcriptional analysis of LN cells of IL-21-treated animals revealed significant inhibition of germinal center (GC) Tfh and B-cell interferon signaling pathways along with enhanced B-cell development and antigen presentation pathways. We conclude that IL-21 treatment at the time of flu vaccination in aging SIV-infected animals modulates the inductive LN GC activity, to reverse SIV-associated LN Tfh and B-cell dysfunction. IL-21 is a potential candidate molecule for immunotherapy to enhance flu vaccine responses in aging PWH who have deficient antibody responses.

Geospatially-resolved public-health surveillance via wastewater sequencing.

Wastewater, which contains everything from pathogens to pollutants, is a geospatially-and temporally-linked microbial fingerprint of a given population. As a result, it can be leveraged for monitoring multiple dimensions of public health across locales and time. Here, we integrate targeted and bulk RNA sequencing (n=1,419 samples) to track the viral, bacterial, and functional content over geospatially distinct areas within Miami Dade County from 2020-2022. First, we used targeted amplicon sequencing (n=966) to track diverse SARS-CoV-2 variants across space and time, and we found a tight correspondence with clinical caseloads from University students (N = 1,503) and Miami-Dade County hospital patients (N = 3,939 patients), as well as an 8-day earlier detection of the Delta variant in wastewater vs. in patients. Additionally, in 453 metatranscriptomic samples, we demonstrate that different wastewater sampling locations have clinically and public-health-relevant microbiota that vary as a function of the size of the human population they represent. Through assembly, alignment-based, and phylogenetic approaches, we also detect multiple clinically important viruses (e.g., norovirus ) and describe geospatial and temporal variation in microbial functional genes that indicate the presence of pollutants. Moreover, we found distinct profiles of antimicrobial resistance (AMR) genes and virulence factors across campus buildings, dorms, and hospitals, with hospital wastewater containing a significant increase in AMR abundance. Overall, this effort lays the groundwork for systematic characterization of wastewater to improve public health decision making and a broad platform to detect emerging pathogens.

Oral and anal microbiome from HIV-exposed individuals: role of host-associated factors in taxa composition and metabolic pathways.

Evidence indicates that the microbiome plays a significant role in HIV immunopathogenesis and associated complications. This study aimed to characterize the oral and anal microbiome of Men who have Sex with Men (MSM) and Transgender Women (TGW), with and without HIV. One hundred and thirty oral and anal DNA-derived samples were obtained from 78 participants and subjected to shotgun metagenomics sequencing for further microbiome analysis. Significant differences in the microbiome composition were found among subjects associated with HIV infection, gender, sex behavior, CD4+ T-cell counts, antiretroviral therapy (ART), and the presence of HPV-associated precancerous anal lesions. Results confirm the occurrence of oncogenic viromes in this high HIV-risk population. The oral microbiome in HIV-associated cases exhibited an enrichment of bacteria associated with periodontal disease pathogenesis. Conversely, anal bacteria showed a significant decrease in HIV-infected subjects (Coprococcus comes, Finegoldia magna, Blautia obeum, Catenibacterium mitsuokai). TGW showed enrichment in species related to sexual transmission, which concurs that most recruited TGW are or have been sex workers. Prevotella bivia and Fusobacterium gonidiaformans were positively associated with anal precancerous lesions among HIV-infected subjects. The enrichment of Holdemanella biformis and C. comes was associated with detectable viral load and ART-untreated patients. Metabolic pathways were distinctly affected by predominant factors linked to sexual behavior or HIV pathogenesis. Gene family analysis identified bacterial gene signatures as potential prognostic and predictive biomarkers for HIV/AIDS-associated malignancies. Conclusions: Identified microbial features at accessible sites are potential biomarkers for predicting precancerous anal lesions and therapeutic targets for HIV immunopathogenesis.

Combined Use of RT-qPCR and NGS for Identification and Surveillance of SARS-CoV-2 Variants of Concern in Residual Clinical Laboratory Samples in Miami-Dade County, Florida.

Over the course of the COVID-19 pandemic, SARS-CoV-2 variants of concern (VOCs) with increased transmissibility and immune escape capabilities, such as Delta and Omicron, have triggered waves of new COVID-19 infections worldwide, and Omicron subvariants continue to represent a global health concern. Tracking the prevalence and dynamics of VOCs has clinical and epidemiological significance and is essential for modeling the progression and evolution of the COVID-19 pandemic. Next generation sequencing (NGS) is recognized as the gold standard for genomic characterization of SARS-CoV-2 variants, but it is labor and cost intensive and not amenable to rapid lineage identification. Here we describe a two-pronged approach for rapid, cost-effective surveillance of SARS-CoV-2 VOCs by combining reverse-transcriptase quantitative polymerase chain reaction (RT-qPCR) and periodic NGS with the ARTIC sequencing method. Variant surveillance by RT-qPCR included the commercially available TaqPath COVID-19 Combo Kit to track S-gene target failure (SGTF) associated with the spike protein deletion H69-V70, as well as two internally designed and validated RT-qPCR assays targeting two N-terminal-domain (NTD) spike gene deletions, NTD156-7 and NTD25-7. The NTD156-7 RT-qPCR assay facilitated tracking of the Delta variant, while the NTD25-7 RT-qPCR assay was used for tracking Omicron variants, including the BA.2, BA.4, and BA.5 lineages. In silico validation of the NTD156-7 and NTD25-7 primers and probes compared with publicly available SARS-CoV-2 genome databases showed low variability in regions corresponding to oligonucleotide binding sites. Similarly, in vitro validation with NGS-confirmed samples showed excellent correlation. RT-qPCR assays allow for near-real-time monitoring of circulating and emerging variants allowing for ongoing surveillance of variant dynamics in a local population. By performing periodic sequencing of variant surveillance by RT-qPCR methods, we were able to provide ongoing validation of the results obtained by RT-qPCR screening. Rapid SARS-CoV-2 variant identification and surveillance by this combined approach served to inform clinical decisions in a timely manner and permitted better utilization of sequencing resources.

Using arts-based research in applied health care: An example from an evaluation of NHS dental contract reform in Wales.

OBJECTIVE: Arts-based research (ABR) refers to the use of art in the research process to help generate, interpret and/or communicate knowledge. We used ABR principles to adapt a centre-staging method to complement a more traditional qualitative approach to evaluate participants' views on dental service reform. METHODS: We asked five individuals in the dental health sector in the National Health Service in Wales to select objects to depict their views on the current reform process and their ideal reform process. This process took place alongside traditional semi-structured interviews with the participants. RESULTS: There were three marked differences in the centre-staging process as compared to the interviews: (1) there was a greater use of symbolism by the participants, (2) the participants put a greater focus on the process of change and (3) the participants were more likely to reveal the emotions underlying their assessments of the reform process. CONCLUSIONS: The arts-based approach adopted appeared to be highly accessible and has the potential to be used in a wide range of applications.

Predicting COVID-19 cases using SARS-CoV-2 RNA in air, surface swab and wastewater samples.

Genomic footprints of pathogens shed by infected individuals can be traced in environmental samples, which can serve as a noninvasive method of infectious disease surveillance. The research evaluates the efficacy of environmental monitoring of SARS-CoV-2 RNA in air, surface swabs and wastewater to predict COVID-19 cases. Using a prospective experimental design, air, surface swabs, and wastewater samples were collected from a college dormitory housing roughly 500 students from March to May 2021 at the University of Miami, Coral Gables, FL. Students were randomly screened for COVID-19 during the study period. SARS-CoV-2 concentration in environmental samples was quantified using Volcano 2nd Generation-qPCR. Descriptive analyses were conducted to examine the associations between time-lagged SARS-CoV-2 in environmental samples and COVID-19 cases. SARS-CoV-2 was detected in air, surface swab and wastewater samples on 52 (63.4 %), 40 (50.0 %) and 57 (68.6 %) days, respectively. On 19 (24 %) of 78 days SARS-CoV-2 was detected in all three sample types. COVID-19 cases were reported on 11 days during the study period and SARS-CoV-2 was also detected two days before the case diagnosis on all 11 (100 %), 9 (81.8 %) and 8 (72.7 %) days in air, surface swab and wastewater samples, respectively. SARS-CoV-2 detection in environmental samples was an indicator of the presence of local COVID-19 cases and a 3-day lead indicator for a potential outbreak at the dormitory building scale. Proactive environmental surveillance of SARS-CoV-2 or other pathogens in multiple environmental media has potential to guide targeted measures to contain and/or mitigate infectious disease outbreaks within communities.

Expanding a Wastewater-Based Surveillance Methodology for DNA Isolation from a Workflow Optimized for SARS-CoV-2 RNA Quantification.

Wastewater-based surveillance (WBS) is a noninvasive, epidemiological strategy for assessing the spread of COVID-19 in communities. This strategy was based upon wastewater RNA measurements of the viral target, severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). The utility of WBS for assessing the spread of COVID-19 has motivated research to measure targets beyond SARS-CoV-2, including pathogens containing DNA. The objective of this study was to establish the necessary steps for isolating DNA from wastewater by modifying a long-standing RNA-specific extraction workflow optimized for SARS-CoV-2 detection. Modifications were made to the sample concentration process and included an evaluation of bead bashing prior to the extraction of either DNA or RNA. Results showed that bead bashing reduced detection of RNA from wastewater but improved recovery of DNA as assessed by quantitative polymerase chain reaction (qPCR). Bead bashing is therefore not recommended for the quantification of RNA viruses using qPCR. Whereas for Mycobacterium bacterial DNA isolation, bead bashing was necessary for improving qPCR quantification. Overall, we recommend 2 separate workflows, one for RNA viruses that does not include bead bashing and one for other microbes that use bead bashing for DNA isolation. The experimentation done here shows that current-standing WBS program methodologies optimized for SARS-CoV-2 need to be modified and reoptimized to allow for alternative pathogens to be readily detected and monitored, expanding its utility as a tool for public health assessment.

Correlative analysis of wastewater trends with clinical cases and hospitalizations through five dominant variant waves of COVID-19.

Wastewater-based epidemiology (WBE) has been utilized to track community infections of Coronavirus Disease 2019 (COVID-19) by detecting RNA of the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), within samples collected from wastewater. The correlations between community infections and wastewater measurements of the RNA can potentially change as SARS-CoV-2 evolves into new variations by mutating. This study analyzed SARS-CoV-2 RNA, and indicators of human waste in wastewater from two sewersheds of different scales (University of Miami (UM) campus and Miami-Dade County Central District wastewater treatment plant (CDWWTP)) during five internally defined COVID-19 variant dominant periods (Initial, Pre-Delta, Delta, Omicron and Post-Omicron wave). SARS-CoV-2 RNA quantities were compared against COVID-19 clinical cases and hospitalizations to evaluate correlations with wastewater SARS-CoV-2 RNA. Although correlations between documented clinical cases and hospitalizations were high, prevalence for a given wastewater SARS-CoV-2 level varied depending upon the variant analyzed. The correlative relationship was significantly steeper (more cases per level found in wastewater) for the Omicron-dominated period. For hospitalization, the relationships were steepest for the Initial wave, followed by the Delta wave with flatter slopes during all other waves. Overall results were interpreted in the context of SARS-CoV-2 virulence and vaccination rates among the community.

Relationships between SARS-CoV-2 in Wastewater and COVID-19 Clinical Cases and Hospitalizations, with and without Normalization against Indicators of Human Waste.

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) in wastewater has been used to track community infections of coronavirus disease-2019 (COVID-19), providing critical information for public health interventions. Since levels in wastewater are dependent upon human inputs, we hypothesize that tracking infections can be improved by normalizing wastewater concentrations against indicators of human waste [Pepper Mild Mottle Virus (PMMoV), ß-2 Microglobulin (B2M), and fecal coliform]. In this study, we analyzed SARS-CoV-2 and indicators of human waste in wastewater from two sewersheds of different scales: a University campus and a wastewater treatment plant. Wastewater data were combined with complementary COVID-19 case tracking to evaluate the efficiency of wastewater surveillance for forecasting new COVID-19 cases and, for the larger scale, hospitalizations. Results show that the normalization of SARS-CoV-2 levels by PMMoV and B2M resulted in improved correlations with COVID-19 cases for campus data using volcano second generation (V2G)-qPCR chemistry (r s = 0.69 without normalization, r s = 0.73 with normalization). Mixed results were obtained for normalization by PMMoV for samples collected at the community scale. Overall benefits from normalizing with measures of human waste depend upon qPCR chemistry and improves with smaller sewershed scale. We recommend further studies that evaluate the efficacy of additional normalization targets.

A pilot study to troubleshoot quality control metrics when assessing circulating miRNA expression data reproducibility across study sites.

BACKGROUND: Given the growing interest in using microRNAs (miRNAs) as biomarkers of early disease, establishment of robust protocols and platforms for miRNA quantification in biological fluids is critical. OBJECTIVE: The goal of this multi-center pilot study was to evaluate the reproducibility of NanoString nCounter™ technology when analyzing the abundance of miRNAs in plasma and cystic fluid from patients with pancreatic lesions. METHODS: Using sample triplicates analyzed across three study sites, we assessed potential sources of variability (RNA isolation, sample processing/ligation, hybridization, and lot-to-lot variability) that may contribute to suboptimal reproducibility of miRNA abundance when using nCounter™, and evaluated expression of positive and negative controls, housekeeping genes, spike-in genes, and miRNAs. RESULTS: Positive controls showed a high correlation across samples from each site (median correlation coefficient, r> 0.9). Most negative control probes had expression levels below background. Housekeeping and spike-in genes each showed a similar distribution of expression and comparable pairwise correlation coefficients of replicate samples across sites. A total of 804 miRNAs showed a similar distribution of pairwise correlation coefficients between replicate samples (p= 0.93). After normalization and selecting miRNAs with expression levels above zero in 80% of samples, 55 miRNAs were identified; heatmap and principal component analysis revealed similar expression patterns and clustering in replicate samples. CONCLUSIONS: Findings from this pilot investigation suggest the nCounter platform can yield reproducible results across study sites. This study underscores the importance of implementing quality control procedures when designing multi-center evaluations of miRNA abundance.

COVID-19 Prediction using Genomic Footprint of SARS-CoV-2 in Air, Surface Swab and Wastewater Samples.

Importance: Genomic footprints of pathogens shed by infected individuals can be traced in environmental samples. Analysis of these samples can be employed for noninvasive surveillance of infectious diseases. Objective: To evaluate the efficacy of environmental surveillance of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) for predicting COVID-19 cases in a college dormitory. Design: Using a prospective experimental design, air, surface swabs, and wastewater samples were collected from a college dormitory from March to May 2021. Students were randomly screened for COVID-19 during the study period. SARS-CoV-2 in environmental samples was concentrated with electronegative filtration and quantified using Volcano 2 nd Generation-qPCR. Descriptive analyses were conducted to examine the associations between time-lagged SARS-CoV-2 in environmental samples and clinically diagnosed COVID-19 cases. Setting: This study was conducted in a residential dormitory at the University of Miami, Coral Gables campus, FL, USA. The dormitory housed about 500 students. Participants: Students from the dormitory were randomly screened, for COVID-19 for 2-3 days / week while entering or exiting the dormitory. Main Outcome: Clinically diagnosed COVID-19 cases were of our main interest. We hypothesized that SARS-CoV-2 detection in environmental samples was an indicator of the presence of local COVID-19 cases in the dormitory, and SARS-CoV-2 can be detected in the environmental samples several days prior to the clinical diagnosis of COVID-19 cases. Results: SARS-CoV-2 genomic footprints were detected in air, surface swab and wastewater samples on 52 (63.4%), 40 (50.0%) and 57 (68.6%) days, respectively, during the study period. On 19 (24%) of 78 days SARS-CoV-2 was detected in all three sample types. Clinically diagnosed COVID-19 cases were reported on 11 days during the study period and SARS-CoV-2 was also detected two days before the case diagnosis on all 11 (100%), 9 (81.8%) and 8 (72.7%) days in air, surface swab and wastewater samples, respectively. Conclusion: Proactive environmental surveillance of SARS-CoV-2 or other pathogens in a community/public setting has potential to guide targeted measures to contain and/or mitigate infectious disease outbreaks. Key Points: Question: How effective is environmental surveillance of SARS-CoV-2 in public places for early detection of COVID-19 cases in a community?Findings: All clinically confirmed COVID-19 cases were predicted with the aid of 2 day lagged SARS-CoV-2 in environmental samples in a college dormitory. However, the prediction efficiency varied by sample type: best prediction by air samples, followed by wastewater and surface swab samples. SARS-CoV-2 was also detected in these samples even on days without any reported cases of COVID-19, suggesting underreporting of COVID-19 cases.Meaning: SARS-CoV-2 can be detected in environmental samples several days prior to clinical reporting of COVID-19 cases. Thus, proactive environmental surveillance of microbiome in public places can serve as a mean for early detection of location-time specific outbreaks of infectious diseases. It can also be used for underreporting of infectious diseases.

Distinct Molecular Signatures of Aging in Healthy and HIV-Infected Individuals.

BACKGROUND: Virally suppressed chronic HIV-infected individuals on antiretroviral therapy experience similar immune impairments as HIV-uninfected elderly. However, they manifest symptoms of premature immune aging such as suboptimal responses to vaccination at a younger age. Mechanisms underlying premature immune aging are unclear. SETTING: The study site was University of Miami Miller School of Medicine. METHODS: In this study, we aimed to identify molecular signatures of aging in HIV-infected (HIV) individuals compared with age-matched healthy control (HC) participants. Transcriptomic profiles of peripheral blood mononuclear cells collected cross-sectionally from study participants were evaluated using RNA sequencing, and genes and pathways associated with age and HIV status were identified and compared between study groups. Generalized linear modeling was used to identify transcriptional signatures associated with age. RESULTS: Despite that fewer differentially expressed genes between young (<40 yrs) and old (>59 yrs) were observed in the HIV group, metabolic and innate immune activation pathways were associated with increasing age in both HIV and HC. Age was also associated with pathways involved with T-cell immune activation in HC and with interferon signaling pathways in HIV. We observed signs of precocious immune aging at the transcriptional level in HIV and defined a transcriptional perturbation associated with innate immunity and glucose metabolism induced by aging in both HC and HIV. CONCLUSION: In this study, we identified distinct molecular signatures predictive of age in HIV versus HC, which suggest precocious immune aging in HIV. Overall, our results highlight the molecular pathways of immune aging in both HC and HIV that may be targeted for additional mechanistic insights or in a therapeutic setting.

Mitochondrial DNA sequence variation and risk of glioma.

BACKGROUND: Malignant gliomas are the most common primary adult brain tumors, with a poor prognosis and ill-defined etiology. Mitochondrial DNA (mtDNA) sequence variation has been linked with certain cancers; however, research on glioma is lacking. METHODS: We examined the association of common (minor allele frequency ≥ 5%) germline mtDNA variants and haplogroups with glioma risk in 1,566 glioma cases and 1,017 controls from a US case-control study, and 425 glioma cases and 1,534 matched controls from the UK Biobank cohort (UKB). DNA samples were genotyped using the UK Biobank array that included a set of common and rare mtDNA variants. Risk associations were examined separately for glioblastoma (GBM) and lower grade tumors (non-GBM). RESULTS: In the US study, haplogroup W was inversely associated with glioma when compared with haplogroup H (OR = 0.43, 95%CI: 0.23-0.79); this association was not demonstrated in the UKB (OR = 1.07, 95%CI: 0.47-2.43). In the UKB, the variant m.3010G > A was significantly associated with GBM (OR = 1.32; 95%CI: 1.01-1.73; p = 0.04), but not non-GBM (1.23; 95%CI: 0.78-1.95; p = 0.38); no similar association was observed in the US study. In the US study, the variant m.14798 T > C, was significantly associated with non-GBM (OR = 0.72; 95%CI: 0.53-0.99), but not GBM (OR = 0.86; 95%CI: 0.66-1.11), whereas in the UKB, a positive association was observed between this variant and GBM (OR = 1.46; 95%CI: 1.06-2.02) but not non-GBM (OR = 0.92; 95%CI: 0.52-1.63). None of these associations were significant after adjustment for multiple testing. CONCLUSION: The association of inherited mtDNA variation, including rare and singleton variants, with glioma risk merits further study.

Comparison of Electronegative Filtration to Magnetic Bead-Based Concentration and V2G-qPCR to RT-qPCR for Quantifying Viral SARS-CoV-2 RNA from Wastewater.

Methods of wastewater concentration (electronegative filtration (ENF) versus magnetic bead-based concentration (MBC)) were compared for the analysis of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), beta-2 microglobulin, and human-coronavirus OC43. Using ENF as the concentration method, two quantitative Polymerase Chain Reaction (qPCR) analytical methods were also compared: Volcano 2nd Generation (V2G)-qPCR and reverse transcriptase (RT)-qPCR measuring three different targets of the virus responsible for the COVID-19 illness (N1, modified N3, and ORF1ab). Correlations between concentration methods were strong and statistically significant for SARS-CoV-2 (r=0.77, p<0.001) and B2M (r=0.77, p<0.001). Comparison of qPCR analytical methods indicate that, on average, each method provided equivalent results with average ratios of 0.96, 0.96 and 1.02 for N3 to N1, N3 to ORF1ab, and N1 to ORF1ab and were supported by significant (p<0.001) correlation coefficients (r =0.67 for V2G (N3) to RT (N1), r =0.74 for V2G (N3) to RT (ORF1ab), r = 0.81 for RT (N1) to RT (ORF1ab)). Overall results suggest that the two concentration methods and qPCR methods provide equivalent results, although variability is observed for individual measurements. Given the equivalency of results, additional advantages and disadvantages, as described in the discussion, are to be considered when choosing an appropriate method.

Planning, commissioning and delivering bespoke short breaks for carers and their partner living with dementia: Challenges and opportunities.

The importance of supporting unpaid carers for people living with dementia (PLWD) is recognised in adult social care policy both nationally and internationally. In the UK, social care legislation emphasises care and support should help people achieve the outcomes that matter to them in their life; this includes the opportunity to take a break from caring routines and responsibilities. Accordingly, there is growing policy and practice interest in short breaks provision to address the diversity of carer break needs and preferences and deliver meaningful outcomes for carers and those they support. This paper reports findings from qualitative staff interviews that offered strategic and operational insights into short breaks provision. It presents a dynamic model of the short break landscape in a region of Wales, describing factors shaping local and regional decision-making. The model identifies key challenges (barriers) and opportunities (enablers) that shape the planning, commissioning and delivery of bespoke short breaks for spousal carers and their partner living with dementia. Through highlighting the interplay between complex context-specific processes and contingences, the model informs initial theory development in short breaks provision.

Mitochondrial DNA sequence variation and risk of meningioma.

BACKGROUND: Risk factors for meningioma include female gender, African American race, high body mass index (BMI), and exposure to ionizing radiation. Although genome-wide association studies (GWAS) have identified two nuclear genome risk loci for meningioma (rs12770228 and rs2686876), the relation between mitochondrial DNA (mtDNA) sequence variants and meningioma is unknown. METHODS: We examined the association of 42 common germline mtDNA variants (minor allele frequency ≥ 5%), haplogroups, and genes with meningioma in 1080 controls and 478 meningioma cases from a case-control study conducted at medical centers in the southeastern United States. Associations were examined separately for meningioma overall and by WHO grade (n = 409 grade I and n = 69 grade II/III). RESULTS: Overall, meningioma was significantly associated with being female (OR 2.85; 95% CI 2.21-3.69), self-reported African American race (OR 2.38, 95% CI 1.41-3.99), and being overweight (OR 1.48; 95% CI 1.11-1.97) or obese (OR 1.70; 95% CI 1.25-2.31). The variant m.16362T > C (rs62581341) in the mitochondrial control region was positively associated with grade II/III meningiomas (OR 2.33; 95% CI 1.14-4.77), but not grade I tumors (OR 0.99; 95% CI 0.64-1.53). Haplogroup L, a marker for African ancestry, was associated with meningioma overall (OR 2.92; 95% CI 1.01-8.44). However, after stratifying by self-reported race, this association was only apparent among the few self-reported Caucasians with this haplogroup (OR 6.35; 95% CI 1.56-25.9). No other mtDNA variant, haplogroup, or gene was associated with meningioma. CONCLUSION: Common mtDNA variants and major mtDNA haplogroups do not appear to have associations with the odds of developing meningioma.

The Effect of JAK1/2 Inhibitors on HIV Reservoir Using Primary Lymphoid Cell Model of HIV Latency.

HIV eradication is hindered by the existence of latent HIV reservoirs in CD4+ T cells. Therapeutic strategies targeting latent cells are required to achieve a functional cure, however the study of latently infected cells from HIV infected persons is extremely challenging due to the lack of biomarkers that uniquely characterize them. In this study, the dual reporter virus HIVGKO was used to investigate latency establishment and maintenance in lymphoid-derived CD4+ T cells. Single cell technologies to evaluate protein expression, host gene expression, and HIV transcript expression were integrated to identify and analyze latently infected cells. FDA-approved, JAK1/2 inhibitors were tested in this system as a potential therapeutic strategy to target the latent reservoir. Latent and productively infected tonsillar CD4+ T cells displayed similar activation profiles as measured by expression of CD69, CD25, and HLADR, however latent cells showed higher CXCR5 expression 3 days post-infection. Single cell analysis revealed a small set of genes, including HIST1-related genes and the inflammatory cytokine, IL32, that were upregulated in latent compared to uninfected and productively infected cells suggesting a role for these molecular pathways in persistent HIV infection. In vitro treatment of HIV-infected CD4+ T cells with physiological concentrations of JAK1/2 inhibitors, ruxolitinib and baricitinib, used in clinical settings to target inflammation, reduced latent and productive infection events when added 24 hr after infection and blocked HIV reactivation from latent cells. Our methods using an established model of HIV latency and lymphoid-derived cells shed light on the biology of latency in a crucial anatomical site for HIV persistence and provides key insights about repurposing baricitinib or ruxolitinib to target the HIV reservoir.

Lessons learned from SARS-CoV-2 measurements in wastewater.

Standardized protocols for wastewater-based surveillance (WBS) for the RNA of SARS-CoV-2, the virus responsible for the current COVID-19 pandemic, are being developed and refined worldwide for early detection of disease outbreaks. We report here on lessons learned from establishing a WBS program for SARS-CoV-2 integrated with a human surveillance program for COVID-19. We have established WBS at three campuses of a university, including student residential dormitories and a hospital that treats COVID-19 patients. Lessons learned from this WBS program address the variability of water quality, new detection technologies, the range of detectable viral loads in wastewater, and the predictive value of integrating environmental and human surveillance data. Data from our WBS program indicated that water quality was statistically different between sewer sampling sites, with more variability observed in wastewater coming from individual buildings compared to clusters of buildings. A new detection technology was developed based upon the use of a novel polymerase called V2G. Detectable levels of SARS-CoV-2 in wastewater varied from 102 to 106 genomic copies (gc) per liter of raw wastewater (L). Integration of environmental and human surveillance data indicate that WBS detection of 100 gc/L of SARS-CoV-2 RNA in wastewater was associated with a positivity rate of 4% as detected by human surveillance in the wastewater catchment area, though confidence intervals were wide (ß ~ 8.99 ∗ ln(100); 95% CI = 0.90-17.08; p < 0.05). Our data also suggest that early detection of COVID-19 surges based on correlations between viral load in wastewater and human disease incidence could benefit by increasing the wastewater sample collection frequency from weekly to daily. Coupling simpler and faster detection technology with more frequent sampling has the potential to improve the predictive potential of using WBS of SARS-CoV-2 for early detection of the onset of COVID-19.

A Non-Coding RNA Network Involved in KSHV Tumorigenesis.

Regulatory pathways involving non-coding RNAs (ncRNAs), such as microRNAs (miRNAs) and long non-coding RNAs (lncRNA), have gained great relevance due to their role in the control of gene expression modulation. Using RNA sequencing of KSHV Bac36 transfected mouse endothelial cells (mECK36) and tumors, we have analyzed the host and viral transcriptome to uncover the role lncRNA-miRNA-mRNA driven networks in KSHV tumorigenesis. The integration of the differentially expressed ncRNAs, with an exhaustive computational analysis of their experimentally supported targets, led us to dissect complex networks integrated by the cancer-related lncRNAs Malat1, Neat1, H19, Meg3, and their associated miRNA-target pairs. These networks would modulate pathways related to KSHV pathogenesis, such as viral carcinogenesis, p53 signaling, RNA surveillance, and cell cycle control. Finally, the ncRNA-mRNA analysis allowed us to develop signatures that can be used to an appropriate identification of druggable gene or networks defining relevant AIDS-KS therapeutic targets.