Novel determinants of intestinal colonization of Salmonella enterica serotype typhimurium identified in bovine enteric infection.

Cattle are naturally infected with Salmonella enterica serotype Typhimurium and exhibit pathological features of enteric salmonellosis that closely resemble those in humans. Cattle are the most relevant model of gastrointestinal disease resulting from nontyphoidal Salmonella infection in an animal with an intact microbiota. We utilized this model to screen a library of targeted single-gene deletion mutants to identify novel genes of Salmonella Typhimurium required for survival during enteric infection. Fifty-four candidate mutants were strongly selected, including numerous mutations in genes known to be important for gastrointestinal survival of salmonellae. Three genes with previously unproven phenotypes in gastrointestinal infection were tested in bovine ligated ileal loops. Two of these mutants, STM3602 and STM3846, recapitulated the phenotype observed in the mutant pool. Complementation experiments successfully reversed the observed phenotypes, directly linking these genes to the colonization defects of the corresponding mutant strains. STM3602 encodes a putative transcriptional regulator that may be involved in phosphonate utilization, and STM3846 encodes a retron reverse transcriptase that produces a unique RNA-DNA hybrid molecule called multicopy single-stranded DNA. The genes identified in this study represent an exciting new class of virulence determinants for further mechanistic study to elucidate the strategies employed by Salmonella to survive within the small intestines of cattle.

Functional connectivity in frontostriatal networks differentiate offspring of parents with substance use disorders from other high-risk youth.

BACKGROUND: Family history (FH) of substance use disorders (SUDs) is known to elevate SUD risk in offspring. However, the influence of FH SUDs has been confounded by the effect of externalizing psychopathologies in the addiction risk neuroimaging literature. Thus, the current study aimed to assess the association between parental SUDs and offspring functional connectivity in samples matched for psychopathology and demographics. METHODS: Ninety 11-12-year-old participants with externalizing disorders were included in the study (48 FH+, 42 FH-). We conducted independent component analyses (ICA) and seed-based analyses (orbitofrontal cortex; OFC, nucleus accumbens (NAcc), dorsolateral prefrontal cortex) with resting state data. RESULTS: FH+ adolescents showed stronger functional connectivity between the right lateral OFC seed and anterior cingulate cortex compared to FH- adolescents (p < 0.05, corrected). Compared to FH-, FH+ adolescents showed stronger negative functional connectivity between the left lateral OFC seed and right postcentral gyrus and between the left NAcc seed and right middle occipital gyrus (p < 0.05, corrected). Poorer emotion regulation was associated with more negative connectivity between right occipital/left NAcc among FH+ adolescents based on the seed-based analysis. FH- adolescents had stronger negative functional connectivity between ventral attention/salience networks and dorsal attention/visuospatial networks in the ICA. CONCLUSIONS: Both analytic methods found group differences in functional connectivity between brain regions associated with executive functioning and regions associated with sensory input (e.g., postcentral gyrus, occipital regions). We speculate that families densely loaded for SUD may confer risk by altered neurocircuitry that is associated with emotion regulation and valuation of external stimuli beyond what would be explained by externalizing psychopathology alone.

Differential Effects of Pergolide and Bromocriptine on Working Memory Performance and Brain Activation after Mild Traumatic Brain Injury.

Dopamine D1 and D2 receptors differ with respect to patterns of regional brain distribution and behavioral effects. Pre-clinical work suggests that D1 agonists enhance working memory, but the absence of selective D1 agonists has constrained using this approach in humans. This study examines working memory performance in mild traumatic brain injury (mTBI) patients when given pergolide, a mixed D1/D2 agonist, compared with bromocriptine, a selective D2 agonist. Fifteen individuals were studied 1 month after mTBI and compared with 17 healthy controls. At separate visits, participants were administered 1.25 mg bromocriptine or 0.05 mg pergolide prior to functional magnetic resonance imaging (MRI) using a working memory task (visual-verbal n-back). Results indicated a significant group-by-drug interaction for mean performance across n-back task conditions, where the mTBI group showed better performance on pergolide relative to bromocriptine, whereas controls showed the opposite pattern. There was also a significant effect of diagnosis, where mTBI patients performed worse than controls, particularly while on bromocriptine, as shown in our prior work. Functional MRI activation during the most challenging task condition (3-back > 0-back contrast) showed a significant group-by-drug interaction, with the mTBI group showing increased activation relative to controls in working memory circuitry while on pergolide, including in the left inferior frontal gyrus. Across participants there was a positive correlation between change in activation in this region and change in performance between drug conditions. Results suggest that activation of the D1 receptor may improve working memory performance after mTBI. This has implications for the development of pharmacological strategies to treat cognitive deficits after mTBI.

Psychosocial impairment following mild blast-induced traumatic brain injury in rats.

Traumatic brain injury (TBI) is associated with increased risk for mental health disorders, impacting post-injury quality of life and societal reintegration. TBI is also associated with deficits in psychosocial processing, defined as the cognitive integration of social and emotional behaviors, however little is known about how these deficits manifest and their contributions to post-TBI mental health. In this pre-clinical investigation using rats, a single mild blast TBI (mbTBI) induced impairment of psychosocial processing in the absence of confounding physical polytrauma, post-injury motor deficits, affective abnormalities, or deficits in non-social behavior. Impairment severity correlated with acute upregulations of a known oxidative stress metabolite, 3-hydroxypropylmercapturic acid (3-HPMA), in urine. Resting state fMRI alterations in the acute post-injury period implicated key brain regions known to regulate psychosocial behavior, including orbitofrontal cortex (OFC), which is congruent with our previous report of elevated acrolein, a marker of neurotrauma and 3-HPMA precursor, in this region following mbTBI. OFC of mbTBI-exposed rats demonstrated elevated mRNA expression of metabotropic glutamate receptors 1 and 5 (mGluR1/5) and injection of mGluR1/5-selective agonist in OFC of uninjured rats approximated mbTBI-induced psychosocial processing impairment, demonstrating a novel role for OFC in this psychosocial behavior. Furthermore, OFC may serve as a hotspot for TBI-induced disruption of psychosocial processing and subsequent mental health disorders.

Identification of novel factors involved in modulating motility of Salmonella enterica serotype typhimurium.

Salmonella enterica serotype Typhimurium can move through liquid using swimming motility, and across a surface by swarming motility. We generated a library of targeted deletion mutants in Salmonella Typhimurium strain ATCC14028, primarily in genes specific to Salmonella, that we have previously described. In the work presented here, we screened each individual mutant from this library for the ability to move away from the site of inoculation on swimming and swarming motility agar. Mutants in genes previously described as important for motility, such as flgF, motA, cheY are do not move away from the site of inoculation on plates in our screens, validating our approach. Mutants in 130 genes, not previously known to be involved in motility, had altered movement of at least one type, 9 mutants were severely impaired for both types of motility, while 33 mutants appeared defective on swimming motility plates but not swarming motility plates, and 49 mutants had reduced ability to move on swarming agar but not swimming agar. Finally, 39 mutants were determined to be hypermotile in at least one of the types of motility tested. Both mutants that appeared non-motile and hypermotile on plates were assayed for expression levels of FliC and FljB on the bacterial surface and many of them had altered levels of these proteins. The phenotypes we report are the first phenotypes ever assigned to 74 of these open reading frames, as they are annotated as 'hypothetical genes' in the Typhimurium genome.

The ABC-type efflux pump MacAB protects Salmonella enterica serovar typhimurium from oxidative stress.

UNLABELLED: Multidrug efflux pumps are integral membrane proteins known to actively excrete antibiotics. The macrolide-specific pump MacAB, the only ABC-type drug efflux pump in Salmonella, has previously been linked to virulence in mice. The molecular mechanism of this link between macAB and infection is unclear. We demonstrate that macAB plays a role in the detoxification of reactive oxygen species (ROS), compounds that salmonellae are exposed to at various stages of infection. macAB is induced upon exposure to H2O2 and is critical for survival of Salmonella enterica serovar Typhimurium in the presence of peroxide. Furthermore, we determined that macAB is required for intracellular replication inside J774.A1 murine macrophages but is not required for survival in ROS-deficient J774.D9 macrophages. macAB mutants also had reduced survival in the intestine in the mouse colitis model, a model characterized by a strong neutrophilic intestinal infiltrate where bacteria may experience the cytotoxic actions of ROS. Using an Amplex red-coupled assay, macAB mutants appear to be unable to induce protection against exogenous H2O2 in vitro, in contrast to the isogenic wild type. In mixed cultures, the presence of the wild-type organism, or media preconditioned by the growth of the wild-type organism, was sufficient to rescue the macAB mutant from peroxide-mediated killing. Our data indicate that the MacAB drug efflux pump has functions beyond resistance to antibiotics and plays a role in the protection of Salmonella against oxidative stress. Intriguingly, our data also suggest the presence of a soluble anti-H2O2 compound secreted by Salmonella cells through a MacAB-dependent mechanism. IMPORTANCE: The ABC-type multidrug efflux pump MacAB is known to be required for Salmonella enterica serovar Typhimurium virulence after oral infection in mice, yet the function of this pump during infection is unknown. We show that this pump is necessary for colonization of niches in infected mice where salmonellae encounter oxidative stress during infection. MacAB is required for growth in cultured macrophages that produce reactive oxygen species (ROS) but is not needed in macrophages that do not generate ROS. In addition, we show that MacAB is required to resist peroxide-mediated killing in vitro and for the inactivation of peroxide in the media. Finally, wild-type organisms, or supernatant from wild-type organisms grown in the presence of peroxide, rescue the growth defect of macAB mutants in H2O2. MacAB appears to participate in the excretion of a compound that induces protection against ROS-mediated killing, revealing a new role for this multidrug efflux pump.