X-ray crystal structures of the pheromone-binding domains of two quorum-hindered transcription factors, YenR of Yersinia enterocolitica and CepR2 of Burkholderia cenocepacia.

The ability of LuxR-type proteins to regulate transcription is controlled by bacterial pheromones, N-acylhomoserine lactones (AHLs). Most LuxR-family proteins require their cognate AHLs for activity, and some of them require AHLs for folding and stability, and for protease-resistance. However, a few members of this family are able to fold, dimerize, bind DNA, and regulate transcription in the absence of AHLs; moreover, these proteins are antagonized by their cognate AHLs. One such protein is YenR of Yersinia enterocolitica, which is antagonized by N-3-oxohexanoyl-l-homoserine lactone (OHHL). This pheromone is produced by the OHHL synthase, a product of the adjacent yenI gene. Another example is CepR2 of Burkholderia cenocepacia, which is antagonized by N-octanoyl-l-homoserine lactone (OHL), whose synthesis is directed by the cepI gene of the same bacterium. Here, we describe the high-resolution crystal structures of the AHL binding domains of YenR and CepR2. YenR was crystallized in the presence and absence of OHHL. While this ligand does not cause large scale changes in the YenR structure, it does alter the orientation of several highly conserved YenR residues within and near the pheromone-binding pocket, which in turn caused a significant movement of a surface-exposed loop.

A genomic investigation of ecological differentiation between free-living and Drosophila-associated bacteria.

Various bacterial taxa have been identified both in association with animals and in the external environment, but the extent to which related bacteria from the two habitat types are ecologically and evolutionarily distinct is largely unknown. This study investigated the scale and pattern of genetic differentiation between bacteria of the family Acetobacteraceae isolated from the guts of Drosophila fruit flies, plant material and industrial fermentations. Genome-scale analysis of the phylogenetic relationships and predicted functions was conducted on 44 Acetobacteraceae isolates, including newly sequenced genomes from 18 isolates from wild and laboratory Drosophila. Isolates from the external environment and Drosophila could not be assigned to distinct phylogenetic groups, nor are their genomes enriched for any different sets of genes or category of predicted gene functions. In contrast, analysis of bacteria from laboratory Drosophila showed they were genetically distinct in their universal capacity to degrade uric acid (a major nitrogenous waste product of Drosophila) and absence of flagellar motility, while these traits vary among wild Drosophila isolates. Analysis of the competitive fitness of Acetobacter discordant for these traits revealed a significant fitness deficit for bacteria that cannot degrade uric acid in culture with Drosophila. We propose that, for wild populations, frequent cycling of Acetobacter between Drosophila and the external environment prevents genetic differentiation by maintaining selection for traits adaptive in both the gut and external habitats. However, laboratory isolates bear the signs of adaptation to persistent association with the Drosophila host under tightly defined environmental conditions.

Convection-enhanced delivery of immunomodulatory therapy for high-grade glioma.

The prognosis for glioblastoma has remained poor despite multimodal standard of care treatment, including temozolomide, radiation, and surgical resection. Further, the addition of immunotherapies, while promising in a number of other solid tumors, has overwhelmingly failed in the treatment of gliomas, in part due to the immunosuppressive microenvironment and poor drug penetrance to the brain. Local delivery of immunomodulatory therapies circumvents some of these challenges and has led to long-term remission in select patients. Many of these approaches utilize convection-enhanced delivery (CED) for immunological drug delivery, allowing high doses to be delivered directly to the brain parenchyma, avoiding systemic toxicity. Here, we review the literature encompassing immunotherapies delivered via CED-from preclinical model systems to clinical trials-and explore how their unique combination elicits an antitumor response by the immune system, decreases toxicity, and improves survival among select high-grade glioma patients.

Agitation Following Severe Traumatic Brain Injury Is a Clinical Sign of Recovery of Consciousness.

Objective: Severe traumatic brain injury (sTBI) often results in disorders of consciousness. Patients emerging from coma frequently exhibit aberrant behaviors such as agitation. These non-purposeful combative behaviors can interfere with medical care. Interestingly, agitation is associated with arousal and is often among the first signs of neurological recovery. A better understanding of these behaviors may shed light on the mechanisms driving the return of consciousness in sTBI patients. This study aims to investigate the association between posttraumatic agitation and the recovery of consciousness. Methods: A retrospective chart review was conducted in 530 adult patients (29.1% female) admitted to Stony Brook University Hospital between January 2011 and December 2019 with a diagnosis of sTBI and Glasgow Coma Scale (GCS) ≤8. Agitation was defined as a Richmond Agitation Sedation Scale (RASS) > +1, or any documentation of equivalently combative and violent behaviors in daily clinical notes. The ability to follow verbal commands was used to define the recovery of consciousness and was assessed daily. Results: Of 530 total sTBI patients, 308 (58.1%) survived. Agitation was present in 169 of all patients and 162 (52.6%) of surviving patients. A total of 273 patients followed commands, and 159 of them developed agitation. Forty patients developed agitation on hospital arrival whereas 119 developed agitation later during their hospital course. Presence of in-hospital agitation positively correlated with command-following (r = 0.315, p < 0.001). The time to develop agitation and time to follow commands showed positive correlation (r = 0.485, p < 0.001). These two events occurred within 3 days in 54 (44.6%) patients, within 7 days in 81 (67.8%) patients, and within 14 days in 96 (80.2%) patients. In 71 (59.7%) patients, agitation developed before command-following; in 36 (30.2%) patients, agitation developed after command-following; in 12 (10.1%) patients, agitation developed on the same day as command-following. Conclusion: Posttraumatic agitation in comatose patients following sTBI is temporally associated with the recovery of consciousness. This behavior indicates the potential for recovery of higher neurological functioning. Further studies are required to identify neural correlates of posttraumatic agitation and recovery of consciousness after sTBI.

Electrocorticography reveals thalamic control of cortical dynamics following traumatic brain injury.

The return of consciousness after traumatic brain injury (TBI) is associated with restoring complex cortical dynamics; however, it is unclear what interactions govern these complex dynamics. Here, we set out to uncover the mechanism underlying the return of consciousness by measuring local field potentials (LFP) using invasive electrophysiological recordings in patients recovering from TBI. We found that injury to the thalamus, and its efferent projections, on MRI were associated with repetitive and low complexity LFP signals from a highly structured phase space, resembling a low-dimensional ring attractor. But why do thalamic injuries in TBI patients result in a cortical attractor? We built a simplified thalamocortical model, which connotes that thalamic input facilitates the formation of cortical ensembles required for the return of cognitive function and the content of consciousness. These observations collectively support the view that thalamic input to the cortex enables rich cortical dynamics associated with consciousness.

Cardiac arrest after severe traumatic brain injury can be survivable with good outcomes.

BACKGROUND: Resuscitation for traumatic cardiac arrest (TCA) in patients with severe traumatic brain injury (sTBI) has historically been considered futile. There is little information on the characteristics and outcomes of these patients to guide intervention and prognosis. The purpose of the current study is to report the clinical characteristics, survival, and long-term neurological outcomes in patients who experienced TCA after sTBI and analyze the factors contributing to survival. METHODS: A retrospective review identified 42 patients with TCA from a total of 402 patients with sTBI (Glasgow Coma Scale (GCS) score ≤8) who were admitted to Stony Brook University Hospital, a level I trauma center, from January 2011 to December 2018. Patient demographics, clinical characteristics, survival, and neurological functioning during hospitalization and at follow-up visits were collected. RESULTS: Of the 42 patients, the average age was 45 years and 21.4% were female. Eight patients survived the injury (19.0%) to discharge and seven survived with good neurological function. Admission GCS score and bilateral pupil reactivity were found to be significant indicators of survival. The mean GCS score was 5.3 in survivors and 3.2 in non-survivors (p=0.020). Age, Injury Severity Score, or cardiac rhythm was not associated with survival. Frequent neuroimaging findings included subarachnoid hemorrhage, subdural hematoma, and diffuse axonal injury. DISCUSSION: TCA after sTBI is survivable and seven out of eight patients in our study recovered with good neurological function. GCS score and pupil reactivity are the best indicators of survival. Our results suggest that due to the possibility of recovery, resuscitation and neurosurgical care should not be withheld from this patient population. LEVEL OF EVIDENCE: Level IV, therapeutic/care management.

Modeling the return to consciousness after severe traumatic brain injury at a large academic level 1 trauma center.

OBJECTIVE: Severe traumatic brain injury (sTBI) carries significant morbidity and mortality. It remains difficult to counsel families on functional prognosis and plan research initiatives aimed at treating traumatic coma. In order to better address these problems, the authors set out to develop statistical models using retrospective data to identify admission characteristics that correlate with time until the return of consciousness, defined as the time to follow commands (TFC). These results were then used to create a TFC score, allowing for rapid identification of patients with predicted prolonged TFC. METHODS: Data were reviewed and collected from medical records of sTBI patients with Glasgow Coma Scale (GCS) motor subscores ≤ 5 who were admitted to Stony Brook University Hospital from January 2011 to July 2018. Data were used to calculate descriptive statistics and build binary logistic regression models to identify admission characteristics that correlated with in-hospital mortality and in-hospital command-following. A Cox proportional hazards model was used to identify admission characteristics that correlated with the length of TFC. A TFC score was developed using the significant variables identified in the Cox regression model. RESULTS: There were 402 adult patients who met the inclusion criteria for this study. The average age was 50.5 years, and 122 (30.3%) patients were women. In-hospital mortality was associated with older age, higher Injury Severity Score (ISS), higher Rotterdam score (head CT grading system), and the presence of bilateral fixed and dilated pupils (p < 0.01). In-hospital command-following was anticorrelated with age, ISS, Rotterdam score, and the presence of a single fixed and dilated pupil (p < 0.05). TFC was anticorrelated with age, ISS, Rotterdam score, and the presence of a single fixed and dilated pupil. Additionally, patients who sustained injuries from falls from standing height had a shorter average TFC. The 3 significant variables from the Cox regression model that explained the most variance were used to create a 4-point TFC score. The most significant of these characteristics were Rotterdam head CT scores, high impact traumas, and the presence of a single fixed and dilated pupil. Importantly, the presence of a single fixed and dilated pupil was correlated with longer TFC but no increase in likelihood of in-hospital mortality. CONCLUSIONS: The creation of the 4-point TFC score will allow clinicians to quickly identify patients with predicted prolonged TFC and estimate the likelihood of command-following at different times after injury. Discussions with family members should take into account the likelihood that patients will return to consciousness and survive after TBI.

In vivo function and comparative genomic analyses of the Drosophila gut microbiota identify candidate symbiosis factors.

Symbiosis is often characterized by co-evolutionary changes in the genomes of the partners involved. An understanding of these changes can provide insight into the nature of the relationship, including the mechanisms that initiate and maintain an association between organisms. In this study we examined the genome sequences of bacteria isolated from the Drosophila melanogaster gut with the objective of identifying genes that are important for function in the host. We compared microbiota isolates with con-specific or closely related bacterial species isolated from non-fly environments. First the phenotype of germ-free Drosophila (axenic flies) was compared to that of flies colonized with specific bacteria (gnotobiotic flies) as a measure of symbiotic function. Non-fly isolates were functionally distinct from bacteria isolated from flies, conferring slower development and an altered nutrient profile in the host, traits known to be microbiota-dependent. Comparative genomic methods were next employed to identify putative symbiosis factors: genes found in bacteria that restore microbiota-dependent traits to gnotobiotic flies, but absent from those that do not. Factors identified include riboflavin synthesis and stress resistance. We also used a phylogenomic approach to identify protein coding genes for which fly-isolate sequences were more similar to each other than to other sequences, reasoning that these genes may have a shared function unique to the fly environment. This method identified genes in Acetobacter species that cluster in two distinct genomic loci: one predicted to be involved in oxidative stress detoxification and another encoding an efflux pump. In summary, we leveraged genomic and in vivo functional comparisons to identify candidate traits that distinguish symbiotic bacteria. These candidates can serve as the basis for further work investigating the genetic requirements of bacteria for function and persistence in the Drosophila gut.