Integrating Oral Health Within Kenyan HIV Research & Policy Structure: Stakeholder Analysis.

Background: Kenya has a remarkably high burden of oral diseases, especially in vulnerable communities like persons with HIV (PWH). In the last few decades, the National AIDS & STI Control Programme has efficiently and successfully provided care and prevention against HIV for people living in Kenya. Objective: To assess the feasibility of integrating oral health into HIV research and policy structures in Kenya. Methods: The study took place between November 2021 and April 2022 in the cities of Nairobi, Kisumu, Mombasa, and Eldoret town. Using a semi-structured interview, three remote and 14 in-person sessions were conducted. Participants included individuals with professional experience in HIV and/or oral health such as researchers, potential mentors, institutional administrators, and other grant-funded experts. A qualitative analysis of recordings was performed by nine pretested independent reviewers, all with qualitative data analysis experience. Areas of interest included research, motivation, obstacles, and support. The free coding phase as well as an iterative grouping analysis (MIRO) was used. Findings: Of the 22 stakeholders interviewed in the study, researchers accounted for the majority (48%) of stakeholders, with the rest composed of practitioners (29%), university administrators (19%), and one public health administrator. University administrators were identified as having the most ability for resource mobilization followed by researchers and practitioners. All participants desired improved health outcomes using an evidence-based approach. The primary motivators were increased networks, collaborations, publications, and bridging the gap between oral health and HIV. While the obstacles to their desires included time and lack of funding, Institutional support through recruitment of qualified personnel, mentors, and mentees was their major desirable support. Conclusion: Stakeholders were unanimous in supporting integrating oral health within the current research and policy environment to address the gap between oral health and HIV, and to improve health outcomes through evidence-based interventions.

Live-cell imaging uncovers the relationship between histone acetylation, transcription initiation, and nucleosome mobility.

Histone acetylation and RNA polymerase II phosphorylation are associated with transcriptionally active chromatin, but their spatiotemporal relationship in live cells remains poorly understood. To address this problem, we combine Fab-based labeling of endogenous protein modifications with single-molecule tracking to quantify the dynamics of chromatin enriched with histone H3 lysine-27 acetylation (H3K27ac) and RNA polymerase II serine-5 phosphorylation (RNAP2-Ser5ph). Our analysis reveals that chromatin enriched with these two modifications is generally separate. In these separated sites, we show that the two modifications are inversely correlated with one another on the minutes time scale and that single nucleosomes within each region display distinct and opposing dynamics on the subsecond time scale. While nucleosomes diffuse ~15% faster in chromatin enriched with H3K27ac, they diffuse ~15% slower in chromatin enriched with RNAP2-Ser5ph. These results argue that high levels of H3K27ac and RNAP2-Ser5ph are not often present together at the same place and time, but rather each marks distinct transcriptionally poised or active sites, respectively.

Gender Differences in Drug Use among Individuals Under Arrest.

Background: Drug monitoring by drug testing of individuals under arrest provides an opportunity to detect drug use patterns within geographic areas. However, women have been omitted from large-scale monitoring efforts in criminal justice populations. The purpose of this study was to examine whether gender differences exist in drug use indicated by oral fluid collected in one U.S. jail. Methods: The study analyzed data collected in 2019-2020 from individuals under arrest (N = 191). Twenty-four percent of the sample identified as female. Oral fluid specimens were collected and then analyzed with enzyme-linked immunosorbent assay and liquid chromatography/tandem mass spectrometry. Logit regression models examined gender differences. Results: Women were more likely to test positive for methamphetamines than men (41% versus 22%, OR = 0.42, 95% CI 0.21-0.84). Significant gender differences were not found for other substances (marijuana, cocaine, and opioids), legality of drugs, or overall drug use. Conclusions: Because the National Institute on Drug Abuse aims to promote health equity, future drug monitoring in criminal justice populations should employ sampling approaches representing both women and men. This research would identify possible gender-based patterns of drug use and inform gender-based policies and clinical practices to prevent and treat drug misuse.

Microbial succession in a marine sediment: Inferring interspecific microbial interactions with marine cable bacteria.

Cable bacteria are long, filamentous, multicellular bacteria that grow in marine sediments and couple sulfide oxidation to oxygen reduction over centimetre-scale distances via long-distance electron transport. Cable bacteria can strongly modify biogeochemical cycling and may affect microbial community networks. Here we examine interspecific interactions with marine cable bacteria (Ca. Electrothrix) by monitoring the succession of 16S rRNA amplicons (DNA and RNA) and cell abundance across depth and time, contrasting sediments with and without cable bacteria growth. In the oxic zone, cable bacteria activity was positively associated with abundant predatory bacteria (Bdellovibrionota, Myxococcota, Bradymonadales), indicating putative predation on cathodic cells. At suboxic depths, cable bacteria activity was positively associated with sulfate-reducing and magnetotactic bacteria, consistent with cable bacteria functioning as ecosystem engineers that modify their local biogeochemical environment, benefitting certain microbes. Cable bacteria activity was negatively associated with chemoautotrophic sulfur-oxidizing Gammaproteobacteria (Thiogranum, Sedimenticola) at oxic depths, suggesting competition, and positively correlated with these taxa at suboxic depths, suggesting syntrophy and/or facilitation. These observations are consistent with chemoautotrophic sulfur oxidizers benefitting from an oxidizing potential imparted by cable bacteria at suboxic depths, possibly by using cable bacteria as acceptors for electrons or electron equivalents, but by an as yet enigmatic mechanism.

Bead Loading Proteins and Nucleic Acids into Adherent Human Cells.

Many live-cell imaging experiments use exogenous particles (e.g., peptides, antibodies, beads) to label or function within cells. However, introducing proteins into a cell across its membrane is difficult. The limited selection of current methods struggles with low efficiency, requires expensive and technically demanding equipment, or functions within narrow parameters. Here, we describe a relatively simple and cost-effective technique for loading DNA, RNA, and proteins into live human cells. Bead loading induces a temporary mechanical disruption to the cell membrane, allowing macromolecules to enter adherent, live mammalian cells. At less than 0.01 USD per experiment, bead loading is the least expensive cell loading method available. Moreover, bead loading does not substantially stress cells or impact their viability or proliferation. This manuscript describes the steps of the bead loading procedure, adaptations, variations, and technical limitations. This methodology is especially suited for live-cell imaging but provides a practical solution for other applications requiring the introduction of proteins, beads, RNA, or plasmids into living, adherent mammalian cells.

Live-cell imaging reveals the spatiotemporal organization of endogenous RNA polymerase II phosphorylation at a single gene.

The carboxyl-terminal domain of RNA polymerase II (RNAP2) is phosphorylated during transcription in eukaryotic cells. While residue-specific phosphorylation has been mapped with exquisite spatial resolution along the 1D genome in a population of fixed cells using immunoprecipitation-based assays, the timing, kinetics, and spatial organization of phosphorylation along a single-copy gene have not yet been measured in living cells. Here, we achieve this by combining multi-color, single-molecule microscopy with fluorescent antibody-based probes that specifically bind to different phosphorylated forms of endogenous RNAP2 in living cells. Applying this methodology to a single-copy HIV-1 reporter gene provides live-cell evidence for heterogeneity in the distribution of RNAP2 along the length of the gene as well as Serine 5 phosphorylated RNAP2 clusters that remain separated in both space and time from nascent mRNA synthesis. Computational models determine that 5 to 40 RNAP2 cluster around the promoter during a typical transcriptional burst, with most phosphorylated at Serine 5 within 6 seconds of arrival and roughly half escaping the promoter in ~1.5 minutes. Taken together, our data provide live-cell support for the notion of efficient transcription clusters that transiently form around promoters and contain high concentrations of RNAP2 phosphorylated at Serine 5.

Microbial Communities Under Distinct Thermal and Geochemical Regimes in Axial and Off-Axis Sediments of Guaymas Basin.

Cold seeps and hydrothermal vents are seafloor habitats fueled by subsurface energy sources. Both habitat types coexist in Guaymas Basin in the Gulf of California, providing an opportunity to compare microbial communities with distinct physiologies adapted to different thermal regimes. Hydrothermally active sites in the southern Guaymas Basin axial valley, and cold seep sites at Octopus Mound, a carbonate mound with abundant methanotrophic cold seep fauna at the Central Seep location on the northern off-axis flanking regions, show consistent geochemical and microbial differences between hot, temperate, cold seep, and background sites. The changing microbial actors include autotrophic and heterotrophic bacterial and archaeal lineages that catalyze sulfur, nitrogen, and methane cycling, organic matter degradation, and hydrocarbon oxidation. Thermal, biogeochemical, and microbiological characteristics of the sampling locations indicate that sediment thermal regime and seep-derived or hydrothermal energy sources structure the microbial communities at the sediment surface.

Surveillance by oral fluid of drugs subject to misuse among individuals under arrest.

Background: Between 1988 and 2013 the U.S. government conducted surveillance of national drug misuse use trends by collecting voluntary urine specimens from individuals under arrest in major counties. It was discontinued for financial reasons. The program was the only national survey that used a bioassay to measure drug use. Other national drug surveys continue to be based entirely on self-reports of drug use.Objective: Given the current opioid and incipient methamphetamine epidemics, this study aimed to demonstrate the feasibility of surveilling drugs subject to misuse among individuals under arrest using oral fluid collected anonymously by jail staff in one U.S. county. This method has never been previously employed with an offender population.Methods: The subjects were adults arrested for any reason and booked in one Midwest county jail in the U.S. between July 2019 - January 2020 (N = 196; 145 males). Oral fluid specimens were provided for research purposes voluntarily and anonymously.Results: 79% of individuals approached consented to participation. The most frequently detected drugs were cannabis (53%), methamphetamine (27%), cocaine (9%) and opioids (11%). Further, 74% tested positive for at least one drug; 36% tested positive for at least one illegal drug, 10% tested positive for at least one possibly illegal drug, and 54% tested positive for at least one legal drug (predominantly cannabis). (Tests for nicotine and ethanol were not included.)Conclusion: The feasibility of collecting oral fluid from individuals under arrest in a jail setting to measure the prevalence of drugs subject to misuse was demonstrated.

mRNA localization is linked to translation regulation in the Caenorhabditis elegans germ lineage.

Caenorhabditis elegans early embryos generate cell-specific transcriptomes despite lacking active transcription, thereby presenting an opportunity to study mechanisms of post-transcriptional regulatory control. We observed that some cell-specific mRNAs accumulate non-homogenously within cells, localizing to membranes, P granules (associated with progenitor germ cells in the P lineage) and P-bodies (associated with RNA processing). The subcellular distribution of transcripts differed in their dependence on 3'UTRs and RNA binding proteins, suggesting diverse regulatory mechanisms. Notably, we found strong but imperfect correlations between low translational status and P granule localization within the progenitor germ lineage. By uncoupling translation from mRNA localization, we untangled a long-standing question: Are mRNAs directed to P granules to be translationally repressed, or do they accumulate there as a consequence of this repression? We found that translational repression preceded P granule localization and could occur independently of it. Further, disruption of translation was sufficient to send homogenously distributed mRNAs to P granules. These results implicate transcriptional repression as a means to deliver essential maternal transcripts to the progenitor germ lineage for later translation.

Horizontal acquisition of a patchwork Calvin cycle by symbiotic and free-living Campylobacterota (formerly Epsilonproteobacteria).

Most autotrophs use the Calvin-Benson-Bassham (CBB) cycle for carbon fixation. In contrast, all currently described autotrophs from the Campylobacterota (previously Epsilonproteobacteria) use the reductive tricarboxylic acid cycle (rTCA) instead. We discovered campylobacterotal epibionts ("Candidatus Thiobarba") of deep-sea mussels that have acquired a complete CBB cycle and may have lost most key genes of the rTCA cycle. Intriguingly, the phylogenies of campylobacterotal CBB cycle genes suggest they were acquired in multiple transfers from Gammaproteobacteria closely related to sulfur-oxidizing endosymbionts associated with the mussels, as well as from Betaproteobacteria. We hypothesize that "Ca. Thiobarba" switched from the rTCA cycle to a fully functional CBB cycle during its evolution, by acquiring genes from multiple sources, including co-occurring symbionts. We also found key CBB cycle genes in free-living Campylobacterota, suggesting that the CBB cycle may be more widespread in this phylum than previously known. Metatranscriptomics and metaproteomics confirmed high expression of CBB cycle genes in mussel-associated "Ca. Thiobarba". Direct stable isotope fingerprinting showed that "Ca. Thiobarba" has typical CBB signatures, suggesting that it uses this cycle for carbon fixation. Our discovery calls into question current assumptions about the distribution of carbon fixation pathways in microbial lineages, and the interpretation of stable isotope measurements in the environment.

Sulfur as an Acceptor to Bromine in Biomolecular Halogen Bonds.

The halogen bond (X-bond) has become an important design element in chemistry, including medicinal chemistry and biomolecular engineering. Although oxygen is the most prevalent and best characterized X-bond acceptor in biomolecules, the interaction is seen with nitrogen, sulfur, and aromatic systems as well. In this study, we characterize the structure and thermodynamics of a Br···S X-bond between a 5-bromouracil base and a phosphorothioate in a model DNA junction. The single-crystal structure of the junction shows the geometry of the Br···S to be variable, while calorimetric studies show that the anionic S acceptor is comparable to or slightly more stable than the analogous O acceptor, with a -3.5 kcal/mol difference in ΔΔH25°C and -0.4 kcal/mol ΔΔG25°C (including an entropic penalty ΔΔS25°C of -10 cal/(mol K)). Thus sulfur is shown to be a favorable acceptor for bromine X-bonds, extending the application of this interaction for the design of inhibitors and biological materials.

Seasonal changes in microbial community structure and activity imply winter production is linked to summer hypoxia in a large lake.

Carbon and nutrient cycles in large temperate lakes such as Lake Erie are primarily driven by phototrophic and heterotrophic microorganisms, although our understanding of these is often constrained to late spring through summer due to logistical constraints. During periods of > 90% ice cover in February of 2008, 2009, and 2010, we collected samples from an icebreaker for an examination of bacterial production as well as microbial community structure. In comparison with summer months (August 2002 and 2010), we tested hypotheses concerning seasonal changes in microbial community diversity and production. Bacterial production estimates were c. 2 orders of magnitude higher (volume normalized) in summer relative to winter. Our observations further demonstrate that the microbial community, including single-celled phototrophs, varied in composition between August and February. Sediment traps deployed and collected over a 3 year period (2008-2011) confirmed that carbon export was ongoing and not limiting winter production. The results support the notion that active primary producers in winter months export carbon to the sediments that is not consumed until the warmer seasons. The establishment of this linkage is a critical observation in efforts to understand the extent and severity of annual summertime formations of a zone of regional hypoxia in Lake Erie.

Plasticity of Total and Intracellular Phosphorus Quotas in Microcystis aeruginosa Cultures and Lake Erie Algal Assemblages.

Blooms of the potentially toxic cyanobacterium Microcystis are common events globally, and as a result significant resources continue to be dedicated to monitoring and controlling these events. Recent studies have shown that a significant proportion of total cell-associated phosphorus (P) in marine phytoplankton can be surface adsorbed; as a result studies completed to date do not accurately report the P demands of these organisms. In this study we measure the total cell-associated and intracellular P as well as growth rates of two toxic strains of Microcystis aeruginosa Kütz grown under a range of P concentrations. The results show that the intracellular P pool in Microcystis represents a percentage of total cell-associated P (50-90%) similar to what has been reported for actively growing algae in marine systems. Intracellular P concentrations (39-147 fg cell(-1)) generally increased with increasing P concentrations in the growth medium, but growth rate and the ratio of total cell-associated to intracellular P remained generally stable. Intracellular P quotas and growth rates in cells grown under the different P treatments illustrate the ability of this organism to successfully respond to changes in ambient P loads, and thus have implications for ecosystem scale productivity models employing P concentrations to predict algal bloom events.

What's in a name? Coming to terms with the child's linguistic environment.

This article reviews the proliferation of terms that have been coined to denote the language environment of the young child. It is argued that terms are often deployed by researchers without due consideration of their appropriateness for particular empirical studies. It is further suggested that just three of the dozen or more available terms meet the needs of child language researchers in most instances: CHILD-DIRECTED SPEECH, INFANT-DIRECTED SPEECH AND EXPOSURE LANGUAGE. The phenomena denoted by these terms are then considered. The term register is generally borrowed for this purpose from sociolinguistics. However, close inspection of this concept reveals that the notion of register needs to be constrained, in specified ways, in order to be of any real value within the field of child language research.

Marine and freshwater cyanophages in a Laurentian Great Lake: evidence from infectivity assays and molecular analyses of g20 genes.

While it is well established that viruses play an important role in the structure of marine microbial food webs, few studies have directly addressed their role in large lake systems. As part of an ongoing study of the microbial ecology of Lake Erie, we have examined the distribution and diversity of viruses in this system. One surprising result has been the pervasive distribution of cyanophages that infect the marine cyanobacterial isolate Synechococcus sp. strain WH7803. Viruses that lytically infect this cyanobacterium were identified throughout the western basin of Lake Erie, as well as in locations within the central and eastern basins. Analyses of the gene encoding the g20 viral capsid assembly protein (a conservative phylogenetic marker for the cyanophage) indicate that these viruses, as well as amplicons from natural populations and the ballast of commercial ships, are related to marine cyanophages but in some cases form a unique clade, leaving questions concerning the native hosts of these viruses. The results suggest that cyanophages may be as important in freshwater systems as they are known to be in marine systems.

Counting-on, trading and partitioning: effects of training and prior knowledge on performance on base-10 tasks.

Factors affecting performance on base-10 tasks were investigated in a series of four studies with a total of 453 children aged 5-7 years. Training in counting-on was found to enhance child performance on base-10 tasks (Studies 2, 3, and 4), while prior knowledge of counting-on (Study 1), trading (Studies 1 and 3), and partitioning (Studies 1 and 4) were associated with enhanced base-10 performance. It emerged that procedural knowledge of counting-on, trading, and partitioning can lead to improvements in procedural knowledge of the base-10 system. The findings lend support to the model of iterative development of conceptual and procedural knowledge advanced by Rittle-Johnson et al. (2001).

Negative input for grammatical errors: effects after a lag of 12 weeks.

Effects of negative input for 13 categories of grammatical error were assessed in a longitudinal study of naturalistic adult-child discourse. Two-hour samples of conversational interaction were obtained at two points in time, separated by a lag of 12 weeks, for 12 children (mean age 2;0 at the start). The data were interpreted within the framework offered by Saxton's (1997, 2000) contrast theory of negative input. Corrective input was associated with subsequent improvements in the grammaticality of child speech for three of the target structures. No effects were found for two forms of positive input: non-contingent models, where the adult produces target structures in non-error-contingent contexts; and contingent models, where grammatical forms follow grammatical child usages. The findings lend support to the view that, in some cases at least, the structure of adult-child discourse yields information on the bounds of grammaticality for the language-learning child.