Extremely acidic proteomes and metabolic flexibility in bacteria and highly diversified archaea thriving in geothermal chaotropic brines.

Few described archaeal, and fewer bacterial, lineages thrive under salt-saturating conditions, such as solar saltern crystallizers (salinity above 30% w/v). They accumulate molar K+ cytoplasmic concentrations to maintain osmotic balance ('salt-in' strategy) and have proteins adaptively enriched in negatively charged acidic amino acids. Here we analysed metagenomes and metagenome-assembled genomes from geothermally influenced hypersaline ecosystems with increasing chaotropicity in the Danakil Depression. Normalized abundances of universal single-copy genes confirmed that haloarchaea and Nanohaloarchaeota encompass 99% of microbial communities in the near-life-limiting conditions of the Western-Canyon Lakes. Danakil metagenome- and metagenome-assembled-genome-inferred proteomes, compared with those of freshwater, seawater and solar saltern ponds up to saturation (6-14-32% salinity), showed that Western-Canyon Lake archaea encode the most acidic proteomes ever observed (median protein isoelectric points ≤4.4). We identified previously undescribed haloarchaeal families as well as an Aenigmatarchaeota family and a bacterial phylum independently adapted to extreme halophily. Despite phylum-level diversity decreasing with increasing salinity-chaotropicity, and unlike in solar salterns, adapted archaea exceedingly diversified in Danakil ecosystems, challenging the notion of decreasing diversity under extreme conditions. Metabolic flexibility to utilize multiple energy and carbon resources generated by local hydrothermalism along feast-and-famine strategies seemingly shapes microbial diversity in these ecosystems near life limits.

Expanded phylogeny of extremely halophilic archaea shows multiple independent adaptations to hypersaline environments.

Extremely halophilic archaea (Haloarchaea, Nanohaloarchaeota, Methanonatronarchaeia and Halarchaeoplasmatales) thrive in saturating salt concentrations where they must maintain osmotic equilibrium with their environment. The evolutionary history of adaptations enabling salt tolerance remains poorly understood, in particular because the phylogeny of several lineages is conflicting. Here we present a resolved phylogeny of extremely halophilic archaea obtained using improved taxon sampling and state-of-the-art phylogenetic approaches designed to cope with the strong compositional biases of their proteomes. We describe two uncultured lineages, Afararchaeaceae and Asbonarchaeaceae, which break the long branches at the base of Haloarchaea and Nanohaloarchaeota, respectively. We obtained 13 metagenome-assembled genomes (MAGs) of these archaea from metagenomes of hypersaline aquatic systems of the Danakil Depression (Ethiopia). Our phylogenomic analyses including these taxa show that at least four independent adaptations to extreme halophily occurred during archaeal evolution. Gene-tree/species-tree reconciliation suggests that gene duplication and horizontal gene transfer played an important role in this process, for example, by spreading key genes (such as those encoding potassium transporters) across extremely halophilic lineages.

System of Psychological Support Based on Positive Suggestions to the Critically Ill Using ICU Doulas.

Surviving critical illness often creates a lasting psychological impact, including depression, anxiety, and post-traumatic stress. Memories of frightening and delusional experiences are the largest potentially modifiable risk factor, but currently, there is no proven intervention to improve these inciting factors. Psychological support based on positive suggestion is a psychotherapeutic approach that can be provided even to patients in altered cognitive states and is therefore a viable psychotherapy intervention throughout the ICU stay. Traditional ICU care team members have limited time and training to provide such psychological support to patients. Doulas are trained supportive companions who have been effectively used to provide patient advocacy and emotional support in other clinical settings and may address this need. Our aim was to train and implement a psychological support based on positive suggestion program for the critically ill using doulas, and measure acceptance of this intervention through stakeholder feedback. METHODS: Doula training included three objectives: an introduction to ICU practice structure and policies; education about fundamental aspects of critical care conditions and procedures; and didactic and hands-on learning experiences in effective use of psychological support based on positive suggestion in the critically ill. Doulas were evaluated at the end of their training and during subsequent clinical activities using competency-based assessments as well as through survey-based questions and interviews with key stakeholders. RESULTS: The ICU doulas performed psychological support based on positive suggestion on 43 critically ill patients in the ICU setting. Stakeholder feedback from nurses, patients, and patient families was positive. The majority (28/32) of surveyed bedside ICU nurses reported that the doulas' involvement was helpful for both patients and nurses alike. All interviewed family members offered positive comments about the ICU doula presence and of the 40 patients who recalled the intervention 37 found it comforting. CONCLUSIONS: Our program successfully trained two doulas to work effectively in the ICU setting performing patient-centered psychological support based on positive suggestion interventions. Their training improved their skill sets and was reported as beneficial by patients, families, and critical care nursing. This training program offers a proof of concept that could be applied in other medical centers, bringing doulas more commonly into the ICU practice to help humanize the experience for patients, families, and medical teams.

CpG-creating mutations are costly in many human viruses.

Mutations can occur throughout the virus genome and may be beneficial, neutral or deleterious. We are interested in mutations that yield a C next to a G, producing CpG sites. CpG sites are rare in eukaryotic and viral genomes. For the eukaryotes, it is thought that CpG sites are rare because they are prone to mutation when methylated. In viruses, we know less about why CpG sites are rare. A previous study in HIV suggested that CpG-creating transition mutations are more costly than similar non-CpG-creating mutations. To determine if this is the case in other viruses, we analyzed the allele frequencies of CpG-creating and non-CpG-creating mutations across various strains, subtypes, and genes of viruses using existing data obtained from Genbank, HIV Databases, and Virus Pathogen Resource. Our results suggest that CpG sites are indeed costly for most viruses. By understanding the cost of CpG sites, we can obtain further insights into the evolution and adaptation of viruses.

Bias in Rating of Rodent Distress during Anesthesia Induction for Anesthesia Compared with Euthanasia.

Selection of an appropriate method of euthanasia involves balancing the wellbeing of the animal during the procedure with the intended use of the animal after death and the physical and psychologic safety of the observer or operator. The recommended practices for anesthesia as compared with euthanasia are very disparate, despite the fact that all chemical methods of euthanasia are anesthetic overdoses. To explain this disparity, this study sought to determine whether perception bias is inherent in the discussion of euthanasia compared with anesthesia. In this study, participants viewed videorecordings of the anesthesia of either 4 rats or 4 mice, from induction to loss of consciousness. Half of the participants were told that they were observing anesthesia; the other half understood that they were observing euthanasia. Participants were asked to rate the distress of the animals by scoring escape behaviors, fear behaviors, respiratory distress, and other distress markers. For mice, the participants generally rated the distress as high when they were told that the mouse was being euthanized, as compared with the participants who were told that the mouse was being anesthetized. For rats, the effect was not as strong, and the distress was generally rated higher when participants were told they were watching anesthesia. Because the interpretation of distress showed bias in both species-even though the bias differed regarding the procedure that interpreted as distressing-this study demonstrates that laboratory animal professionals must consider the influence of potential perception bias when developing policies for euthanasia and anesthesia.