Perception and Correlates of Opioid Overdose Risk Among Overdose Survivors Who Use Nonprescribed Opioids in San Francisco and Boston.

BACKGROUND: Understanding opioid overdose risk perception may inform overdose prevention strategies. METHODS: We used baseline data from a randomized overdose prevention trial, in San Francisco, CA, and Boston, MA, among people who used nonprescribed opioids, survived an overdose in the past 3 years, and had received naloxone. Participants were asked how likely they were to overdose in the next 4 months. We combined "extremely likely" and "likely" (higher risk perception) and "neutral," "unlikely," and "extremely unlikely" (lower risk perception). We performed bivariate analyses and separate multivariable logistic regression models of risk perception across (1) sociodemographic, (2) substance use, and (3) overdose risk behavior measures. Covariates were selected a priori or significant in bivariate analyses. RESULTS: Among 268 participants, 88% reported at least 1 overdose risk behavior; however, only 21% reported higher risk perception. The adjusted odds ratio (AOR) of higher risk perception was 2.41 (95% confidence interval [CI]: 1.10-5.30) among those unhoused in the past 4 months, 2.06 (95% CI: 1.05-4.05) among those using opioids in a new place, and 5.61 (95% CI: 2.82-11.16) among those who had overdosed in the past 4 months. Living in Boston was associated with higher risk perception in all 3 models (AOR = 2.00-2.46, 95% CI: 1.04-4.88). CONCLUSIONS: Despite prevalent risk behaviors, a minority of participants perceived themselves to be at higher risk of overdose. Nonetheless, some known risk factors for overdose were appropriately associated with risk perception. Fentanyl has been prevalent in Boston for longer than San Francisco, which may explain the higher risk perception there.

Getting a handle on rat familiarization: The impact of handling protocols on classic tests of stress in Rattus norvegicus.

Experimenter familiarization with laboratory rodents through handling prior to experimentation is an important practice in neurobehavioral research and is implicated in stress, study variability, and replicability. Unfortunately, different handling protocols have not been thoroughly examined. Determining optimal experimenter familiarization protocols is expected to reduce animal stress and thus improve welfare and data consistency. The impact of different handling protocols was determined through behavioral assessments (i.e. elevated plus maze, light/dark box, open field) as well as via analysis of fecal boli counts, ultrasonic vocalizations, and blood corticosterone. Male and female Sprague Dawley rats were distributed among three groups: never handled, picked-up, and handled for 5 min once daily over five days. Handled and picked-up rats spent more time in open arms and less time in closed arms of the elevated plus maze and more time in the center and less time at the perimeter of the open field compared to rats that were never handled, indicating that handled and picked-up rats were less anxious than those that were never handled. Male rats consistently defecated more frequently throughout the handling process and throughout behavioral testing, whereas females showed greater concentrations of blood corticosterone. Female rats were found to emit more 50-kHz calls and fewer 22-kHz calls compared to males. The results observed suggest that picking animals up may suffice as a handling method compared to time-intensive handling procedures, and that there are significant sex differences in response to handling.

Microencapsulation and in situ incubation methodology for the cultivation of marine bacteria.

Environmental microorganisms are important sources of biotechnology innovations; however, the discovery process is hampered by the inability to culture the overwhelming majority of microbes. To drive the discovery of new biotechnology products from previously unculturable microbes, several methods such as modification of media composition, incubation conditions, single-cell isolation, and in situ incubation, have been employed to improve microbial recovery from environmental samples. To improve microbial recovery, we examined the effect of microencapsulation followed by in situ incubation on the abundance, viability, and diversity of bacteria recovered from marine sediment. Bacteria from marine sediment samples were resuspended or encapsulated in agarose and half of each sample was directly plated on agar and the other half inserted into modified Slyde-A-Lyzer™ dialysis cassettes. The cassettes were incubated in their natural environment (in situ) for a week, after which they were retrieved, and the contents plated. Colony counts indicated that bacterial abundance increased during in situ incubation and that cell density was significantly higher in cassettes containing non-encapsulated sediment bacteria. Assessment of viability indicated that a higher proportion of cells in encapsulated samples were viable at the end of the incubation period, suggesting that agarose encapsulation promoted higher cell viability during in situ incubation. One hundred and 46 isolates were purified from the study (32-38 from each treatment) to assess the effect of the four treatments on cultivable bacterial diversity. In total, 58 operational taxonomic units (OTUs) were identified using a 99% 16S rRNA gene sequence identity threshold. The results indicated that encapsulation recovered greater bacterial diversity from the sediment than simple resuspension (41 vs. 31 OTUs, respectively). While the cultivable bacterial diversity decreased by 43%-48% after in situ incubation, difficult-to-culture (Verrucomicrobia) and obligate marine (Pseudoalteromonas) taxa were only recovered after in situ incubation. These results suggest that agarose encapsulation coupled with in situ incubation in commercially available, low-cost, diffusion chambers facilitates the cultivation and improved recovery of bacteria from marine sediments. This study provides another tool that microbiologists can use to access microbial dark matter for environmental, biotechnology bioprospecting.

The Effects of Rearing in a Shelved Environment on Behavioral and Physiological Markers of Welfare in Rats (Rattus norvegicus).

Early-life experiences are critical modifiers of development. An important component of early-life experience is the nature of maternal interactions, which can be modified by stress. During rearing, mothers are typically allocated to single-level cages where they are readily accessible to pups, a potentially stressful scenario not reflective of nature. Accordingly, mothers regularly removed from the rearing environment interact differently with their offspring, leading to long-term changes in offspring physiology and behavior. Such changes commonly include modifications within the hypothalamic-pituitary-adrenal axis, of which corticosterone is a major component. Modifications in the hypothalamic-pituitary-adrenal axis may also be manifested through changes in affective behavior and assessed via tests such as the open field and elevated plus maze as well as via ultrasonic vocalization (USV) analysis. As a means of assessing the impact of rearing in a shelved environment, we allocated mothers to standard single-level cages or cages with an integrated shelf, which allowed the mother to temporarily escape pups. While there were no differences in fecal cortico-sterone, behavior in the elevated plus maze, or USVs, male rats reared in standard cages weighed more, and all standard single-level housed rats spent more time in the center of the open field. The observed differences indicate that allocating nursing mothers to shelved environments throughout the postnatal period has long-lasting effects on offspring behavior that must be considered when establishing dam enrichment protocols.

Effects of Matrix Composition and Temperature on Viability and Metabolic Activity of Microencapsulated Marine Bacteria.

To enhance the discovery of novel natural products, various innovations have been developed to aid in the cultivation of previously unculturable microbial species. One approach involving the microencapsulation of bacteria has been gaining popularity as a new cultivation technique, with promising applications. Previous studies demonstrated the success of bacterial encapsulation; however, they highlighted that a key limitation of encapsulating bacteria within agarose is the high temperature required for encapsulation. Encapsulation of bacteria within agarose typically requires a temperature high enough to maintain the flow of agarose through microfluidic devices without premature gelation. Given the sensitivity of many bacterial taxa to temperature, the effect of various agarose-based encapsulating matrices on marine bacterial viability was assessed to further develop this approach to bacterial culture. It was determined that lowering the temperature of encapsulation via the use of low-gelling-temperature agarose, as well as the addition of nutrients to the matrix, significantly improved the viability of representative marine sediment bacteria in terms of abundance and metabolic activity. Based on these findings, the use of low-gelling-temperature agarose with supplemental nutrients is recommended for the encapsulation of marine bacteria obtained from temperate habitats.

Physical Presence during Moral Action in Immersive Virtual Reality.

Research on morality has focused on differences in moral judgment and action. In this study, we investigated self-reported moral reasoning after a hypothetical moral dilemma was presented on paper, and moral reasoning after that very same dilemma was experienced in immersive virtual reality (IVR). We asked open-ended questions and used content analysis to determine moral reasoning in a sample of 107 participants. We found that participants referred significantly more often to abstract principles and consequences for themselves (i.e., it is against the law) after the paper-based moral dilemma compared to the IVR dilemma. In IVR participants significantly more often referred to the consequences for the people involved in the dilemma (i.e., not wanting to hurt that particular person). This supports the separate process theory, suggesting that decision and action might be different moral concepts with different foci regarding moral reasoning. Using simulated moral scenarios thus seems essential as it illustrates possible mechanisms of empathy and altruism being more relevant for moral actions especially given the physical presence of virtual humans in IVR.

Development of a microbe domestication pod (MD Pod) for in situ cultivation of micro-encapsulated marine bacteria.

Microbial marine natural products hold significant potential for the discovery of new bioactive therapeutics such as antibiotics. Unfortunately, this discovery is hindered by the inability to culture the majority of microbes using traditional laboratory approaches. While many new methods have been developed to increase cultivability, a high-throughput in situ incubation chamber capable of simultaneously isolating individual microbes while allowing cellular communication has not previously been reported. Development of such a device would expedite the discovery of new microbial taxa and, thus, facilitate access to their associated natural products. In this study, this concept is achieved by the development of a new device termed by the authors as the microbe domestication (MD) Pod. The MD Pod enables single-cell cultivation by isolating marine bacterial cells in agarose microbeads produced using microfluidics, while allowing potential transmission of chemical signals between cells during in situ incubation in a chamber, or "Pod," that is deployed in the environment. The design of the MD Pod was optimized to ensure the use of biocompatible materials, allow for simple assembly in a field setting, and maintain sterility throughout incubation. The encapsulation process was designed to ensure that the viability of marine sediment bacteria was not adversely impacted by the encapsulation process. The process was validated using representative bacteria isolated from temperate marine sediment samples: Marinomonas polaris, Psychrobacter aquimaris, and Bacillus licheniformis. The overall process appeared to promote metabolic activity of most representative species. Thus, microfluidic encapsulation of marine bacteria and subsequent in situ incubation in the MD Pod is expected to accelerate marine natural products discovery by increasing the cultivability of marine bacteria.

Social stress shortens lifespan in mice.

Stress and low socioeconomic status in humans confer increased vulnerability to morbidity and mortality. However, this association is not mechanistically understood nor has its causation been explored in animal models thus far. Recently, cellular senescence has been suggested as a potential mechanism linking lifelong stress to age-related diseases and shorter life expectancy in humans. Here, we established a causal role for lifelong social stress on shortening lifespan and increasing the risk of cardiovascular disease in mice. Specifically, we developed a lifelong chronic psychosocial stress model in which male mouse aggressive behavior is used to study the impact of negative social confrontations on healthspan and lifespan. C57BL/6J mice identified through unbiased cluster analysis for receiving high while exhibiting low aggression, or identified as subordinate based on an ethologic criterion, had lower median and maximal lifespan, and developed earlier onset of several organ pathologies in the presence of a cellular senescence signature. Critically, subordinate mice developed spontaneous early-stage atherosclerotic lesions of the aortic sinuses characterized by significant immune cells infiltration and sporadic rupture and calcification, none of which was found in dominant subjects. In conclusion, we present here the first rodent model to study and mechanistically dissect the impact of chronic stress on lifespan and disease of aging. These data highlight a conserved role for social stress and low social status on shortening lifespan and increasing the risk of cardiovascular disease in mammals and identify a potential mechanistic link for this complex phenomenon.

Infant Oral Mutilation.

Ebinyo refers to the practice of removing primary canine tooth follicles in infants without anesthetic by African traditional healers or elders using unsterilized instruments. This report describes a case of ebinyo or infant oral mutilation (IOM) and associated sequelae in a child adopted from a remote African tribe. The intraoral examination revealed that the patient was missing his primary maxillary and mandibular canines. The maxillary anterior periapical radiograph displayed a dysmorphic ectopic unerupted maxillary right primary canine positioned mesial to the maxillary right primary first molar. Periapical films taken confirmed partial or complete absence of the patient's primary mandibular left (73) and mandibular right (83) canines, and a bitewing and periapical film confirmed the absence of the patient's primary maxillary left (63) canine. The permanent canines will be monitored for possible hypoplasia secondary to trauma to the tooth buds during extirpation of the primary canines. Research presented in this report reveals that there are serious health implications involved with the practice of ebinyo.

Imprinting defects at human 14q32 locus alters gene expression and is associated with the pathobiology of osteosarcoma.

Osteosarcoma is the most common primary bone malignancy affecting children and adolescents. Although several genetic predisposing conditions have been associated with osteosarcoma, our understanding of its pathobiology is rather limited. Here we show that, first, an imprinting defect at human 14q32-locus is highly prevalent (87%) and specifically associated with osteosarcoma patients < 30 years of age. Second, the average demethylation at differentially methylated regions (DMRs) in the 14q32-locus varied significantly compared to genome-wide demethylation. Third, the 14q32-locus was enriched in both H3K4-me3 and H3K27-me3 histone modifications that affected expression of all imprinted genes and miRNAs in this region. Fourth, imprinting defects at 14q32 - DMRs are present in triad DNA samples from affected children and their biological parents. Finally, imprinting defects at 14q32-DMRs were also observed at higher frequencies in an Rb1/Trp53 mutation-induced osteosarcoma mouse model. Further analysis of normal and tumor tissues from a Sleeping Beauty mouse model of spontaneous osteosarcoma supported the notion that these imprinting defects may be a key factor in osteosarcoma pathobiology. In conclusion, we demonstrate that imprinting defects at the 14q32 locus significantly alter gene expression, may contribute to the pathogenesis of osteosarcoma, and could be predictive of survival outcomes.

Isotope composition and volume of Earth's early oceans.

Oxygen and hydrogen isotope compositions of Earth's seawater are controlled by volatile fluxes among mantle, lithospheric (oceanic and continental crust), and atmospheric reservoirs. Throughout geologic time the oxygen mass budget was likely conserved within these Earth system reservoirs, but hydrogen's was not, as it can escape to space. Isotopic properties of serpentine from the approximately 3.8 Ga Isua Supracrustal Belt in West Greenland are used to characterize hydrogen and oxygen isotope compositions of ancient seawater. Archaean oceans were depleted in deuterium [expressed as δD relative to Vienna standard mean ocean water (VSMOW)] by at most 25 ± 5‰, but oxygen isotope ratios were comparable to modern oceans. Mass balance of the global hydrogen budget constrains the contribution of continental growth and planetary hydrogen loss to the secular evolution of hydrogen isotope ratios in Earth's oceans. Our calculations predict that the oceans of early Earth were up to 26% more voluminous, and atmospheric CH(4) and CO(2) concentrations determined from limits on hydrogen escape to space are consistent with clement conditions on Archaean Earth.

Serpentinite and the dawn of life.

Submarine hydrothermal vents above serpentinite produce chemical potential gradients of aqueous and ionic hydrogen, thus providing a very attractive venue for the origin of life. This environment was most favourable before Earth's massive CO(2) atmosphere was subducted into the mantle, which occurred tens to approximately 100 Myr after the moon-forming impact; thermophile to clement conditions persisted for several million years while atmospheric pCO(2) dropped from approximately 25 bar to below 1 bar. The ocean was weakly acid (pH ∼ 6), and a large pH gradient existed for nascent life with pH 9-11 fluids venting from serpentinite on the seafloor. Total CO(2) in water was significant so the vent environment was not carbon limited. Biologically important phosphate and Fe(II) were somewhat soluble during this period, which occurred well before the earliest record of preserved surface rocks approximately 3.8 billion years ago (Ga) when photosynthetic life teemed on the Earth and the oceanic pH was the modern value of approximately 8. Serpentinite existed by 3.9 Ga, but older rocks that might retain evidence of its presence have not been found. Earth's sequesters extensive evidence of Archaean and younger subducted biological material, but has yet to be exploited for the Hadean record.