Persistent Phenotype Research Articles

Human epididymis protein 4-annexin II binding promotes aberrant epithelial-fibroblast crosstalk in pulmonary fibrosis

Invasive lung myofibroblasts are the main cause of tissue remodeling in idiopathic pulmonary fibrosis (IPF). A key mechanism contributing to this important feature is aberrant crosstalk between the abnormal/injured lung epithelium and pulmonary fibroblasts. Here, we demonstrate that lungs from patients with IPF and from mice with bleomycin (BLM)-induced pulmonary fibrosis (PF) are characterized by the induction of human epididymis protein 4 (HE4) overexpression in epithelial cells. HE4 knockdown primarily in epithelial cells attenuates BLM-induced PF in mice, whereas the administration of recombinant mouse HE4 exacerbates fibrosis after BLM stimulation. Mechanistic analysis shows that HE4 and annexin II (ANXA2) specific binding enhances the profibrotic phenotype in epithelial cells, and directly promotes lung fibroblast activation, leading to aberrant epithelial-fibroblast crosstalk and the persistent myofibroblast phenotype. The HE4 and ANXA2 binding site is located after the 30th amino acid at the N terminus of the HE4 molecule. Finally, intratracheal administration of HE4 shRNA lentivirus protects mice against BLM-induced PF. These data suggest that HE4 can serve as a potential therapeutic target in the treatment of IPF.

Valproate-Induced Model of Autism in Adult Zebrafish: A Systematic Review.

Autism spectrum disorder (ASD) is a neurodevelopmental disorder characterized by deficits in social skills and the presence of repetitive and restricted behaviors and interests. The social behavior of the zebrafish (Danio rerio) makes this organism a valuable tool for modeling ASD in order to explore the social impairment typical of this disorder. In addition to transgenic models, exposure of zebrafish embryos to valproic acid (VPA) has been found to produce ASD-like symptoms. This review first sets out to examine the existing literature on adult social behavior in the zebrafish VPA-induced model of autism, and the authors also aim to identify the ideal VPA dosage able to induce a persistent and long-lasting ASD-like phenotype while minimizing the suffering and distress of research animals in compliance with the principles of replacement, refinement, and reduction (3Rs).

Phage-Encoded Virulence Factor, Gp05, Alters Membrane Phospholipids and Reduces Antimicrobial Susceptibility in Methicillin-Resistant Staphylococcus aureus.

Methicillin-resistant Staphylococcus aureus (MRSA) is a leading pathogen causing severe endovascular infections. The prophage-encoded protein Gp05 has been identified as a critical virulence factor that contributes to MRSA persistence during vancomycin (VAN) treatment in an experimental endocarditis model. However, the underlining mechanisms driving this persistence phenotype remain poorly understood. The current study aimed to elucidate the genetic factors contributing to Gp05-associated MRSA persistence by utilizing RNA sequencing (RNA-seq) on an isogenic MRSA strain set, including a clinical persistent bacteremia isolate (PB 300-169), its isogenic chromosomal gp05 deletion mutant, and gp05-complemented strains. RNA-seq analysis revealed significantly downregulation of the graSR-vraFG regulatory system and its downstream genes, mprF and dltABCD, in the gp05 deletion mutant compared to the wild-type and gp05-complemented strains. Notably, this downregulation led to a substantial shift in cell membrane composition, with a marked increase in negatively charged phosphatidylglycerol (PG) and a concomitant decrease in positively charged lysyl-PG (LPG). These changes in membrane lipid composition resulted in increased susceptibility of the gp05 deletion mutant to human cationic antimicrobial peptide (CAMP) LL-37, polymorphonuclear neutrophil (PMN) and VAN. Similar findings were observed in an isogenic gp05 overexpression strain set with different genetic background (MRSA USA300 JE2). These findings suggest that Gp05 plays a pivotal role in MRSA persistence by modulating cell surface components and surface charge. This study provides new insights into the molecular mechanisms underlying Gp05-mediated persistence in MRSA endovascular infections and highlights potential therapeutic targets to combat persistent MRSA infections.

The Homeobox Transcription Factor Cux1 Coordinates Postnatal Epithelial Developmental Timing but Is Dispensable for Lung Organogenesis and Regeneration.

Lung epithelial progenitors use a complex network of known and predicted transcriptional regulators to influence early lung development. Here, we evaluate the function of one predicted regulator, Cux1, that we identified from transcriptional regulatory analysis of the SOX9+ distal lung progenitor network. We generated a new Cux1-floxed mouse model and created an epithelial-specific knockout of Cux1 using Shh-Cre (Cux1ShhCre-LOF). Postnatal Cux1ShhCre-LOF animals recapitulated key skin phenotypic features found in prior constitutive Cux1 knockout animals, confirming functionality of our new floxed model. Postnatal Cux1ShhCre-LOF mice displayed subtle alveolar simplification and a transient delay in alveologenesis and alveolar type 1 cell development without persistent lung phenotypes. Cux1ShhCre-LOF mice developed failure to thrive in their second and third weeks of life due to delayed ileal maturation, which similarly resolves by postnatal day 35. Finally, we challenged Cux1ShhCre-LOF with influenza-mediated lung injury to demonstrate that Cux1ShhCre-LOF mice undergo productive alveolar regeneration that is indistinguishable from WT animals. Together, these findings indicate that epithelial-specific loss of Cux1 leads to transient developmental delays in the skin, lung, and intestine without defects in definitive organogenesis. We conclude that Cux1 function is required for temporal optimization of developmental maturation in multiple organs with implications for susceptibility windows in developmental disease pathogenesis.

Developmental trajectories of atopic dermatitis with multi-omics approaches in the infant gut: COCOA birth cohort

BackgroundAn understanding of the phenotypes and endotypes of atopic dermatitis (AD) is essential for developing precision therapies. Recent studies have demonstrated evidence for the gut-skin axis in AD. ObjectiveTo determine the natural course and clinical characteristics of AD phenotypes and investigate their mechanisms based on multi-omics analyses. MethodsLatent class trajectory analysis was used to AD phenotype in 2247 children who were followed until 9 years of age from the COhort for Childhood Origin of Asthma and allergic diseases (COCOA) birth cohort study. Multi-omics analyses (microbiome, metabolites, and gut epithelial cell transcriptome) using stool samples collected at 6 months of age were performed to elucidate the underlying mechanisms of AD phenotypes. ResultsFive AD phenotypes were classified as follows: never/infrequent, early-onset transient, intermediate-transient, late-onset, and early-onset persistent. Early-onset persistent and late-onset phenotypes showed increased risks of food allergy and wheezing treatment ever, with bronchial hyperresponsiveness only evident in the early-onset persistent phenotype. Multi-omics analyses revealed a significantly lower relative abundance of Ruminococcus gnavus and a decreased gut acetate level in the early-onset persistent phenotype, with potential associations to ACSS2, JAK-STAT signaling, and systemic Th2 inflammation. The early-onset transient phenotype was associated with AMPK and/or chemokine signaling regulation, whereas the late-onset phenotype was linked with IL-17 and barrier dysfunction. ConclusionsMulti-omics profiling in early life may offer insights into different mechanisms underlying AD phenotypes in children.

Race for the surface between THP-1 macrophages and Staphylococcus aureus on various titanium implants with well-defined topography and wettability.

Gristina et al. (1987) suggested the fate of a biomaterial is decided in a “race for the surface” between pathogens and the host. To gain deeper insight into the mechanisms behind this concept, we investigated the “race for the surface” across three co-culture scenarios with THP-1 macrophages and Staphylococcus aureus (1:1 ratio), varying the order of addition: (i) simultaneous, (ii) macrophages first, and (iii) S. aureus first, on six Ti6Al4V-ELI surfaces modified with specific topographies and wettability. The outcome of the race for the surface was not influenced by these biomaterials but by the chronological introduction of macrophages and S. aureus. When macrophages and S. aureus arrived simultaneously, macrophages won the race, leading to the lowest number of viable S. aureus through rapid phagocytosis and killing. When macrophages arrived and established first, macrophages still prevailed but under greater challenge resulting from the lower bacterial killing efficiency of adherent macrophages and numerous viable intracellular bacteria, supporting the concept of the so-called immunocompromised zone around implants (upregulation of TLR-2 receptor and pro-inflammatory IL-1β). When S. aureus arrived first establishing a biofilm, bacteria won the race, leading to macrophage dysfunction and cell death (upregulation of FcγR and TLR-2 receptors, NF-κB signaling, NOX2 mediated reactive oxygen species), contributing to a persistent biofilm phenotype (upregulation of clfA, icaA, sarA, downregulation of agrA, hld, lukAB) and intracellular survival of S. aureus (lipA upregulation). The clinical implications are bacterial colonization of the implant and persistence of intracellular bacteria in periprosthetic tissues, which can lead to infection chronicity. Statement of SignificanceGristina et al. (1987) suggested the fate of a biomaterial is decided in a “race for the surface” between bacterial pathogens and host cells. There is a lack of in vitro co-culture models and knowledge on macrophage-S. aureus interactions on biomaterial surfaces, and none has evaluated the expression of virulence factors in S. aureus biofilms.We have successfully developed co-culture models and molecular panels and elucidated important cellular and molecular interactions between macrophages and S. aureus on a broad range of titanium biomaterials with well-defined surface topography and wettability. Our findings highlight the critical role of biofilm formation and the chronological order of bacteria or macrophage arrival in determining the fate of the surface.

Carcinogenic Mechanisms of Hexavalent Chromium: From DNA Breaks to Chromosome Instability and Neoplastic Transformation.

Hexavalent chromium [Cr(VI)] is a well-established human carcinogen, yet the mechanisms by which it leads to carcinogenic outcomes is still unclear. As a driving factor in its carcinogenic mechanism, Cr(VI) causes DNA double strand breaks and break-repair deficiency, leading to the development of chromosome instability. Therefore, the aim of this review is to discuss studies assessing Cr(VI)-induced DNA double strand breaks, chromosome damage and instability, and neoplastic transformation including cell culture, experimental animal, human pathology and epidemiology studies. Recent findings confirm Cr(VI) induces DNA double strand breaks, chromosome instability and neoplastic transformation in exposed cells, animals and humans, emphasizing these outcomes as key steps in the mechanism of Cr(VI) carcinogenesis. Moreover, recent findings suggest chromosome instability is a key phenotype in Cr(VI)-neoplastically transformed clones and is an inheritable and persistent phenotype in exposed cells, once more suggesting chromosome instability as central in the carcinogenic mechanism. Although limited, some studies have demonstrated DNA damage and epigenetic modulation are also key outcomes in biopsies from chromate workers that developed lung cancer. Additionally, we also summarized new studies showing Cr(VI) causes genotoxic and clastogenic effects in cells from wildlife, such as sea turtles, whales, and alligators. Overall, across the literature, it is clear that Cr(VI) causes neoplastic transformation and lung cancer. Many studies measured Cr(VI)-induced increases in DNA double strand breaks, the most lethal type of breaks clearly showing that Cr(VI) is genotoxic. Unrepaired or inaccurately repaired breaks lead to the development of chromosome instability, which is a common phenotype in Cr(VI) exposed cells, animals, and humans. Indeed, many studies show Cr(VI) induces both structural and numerical chromosome instability. Overall, the large body of literature strongly supports the conclusion that Cr(VI) causes DNA double strand breaks, inhibits DNA repair and chromosome instability, which are key to the development of Cr(VI)-induced cell transformation.

Severe aortic stenosis is associated with left atrial prothrombotic flow changes that persists despite successful aortic valve replacement

Abstract Background Severe aortic stenosis (AS) is associated with ischaemic stroke independently of atrial fibrillation (AF) or other causes. Even after aortic valve replacement (AVR), patients remain affected by up to 15% five-year rate of incident ischaemic strokes independently of AVR-related complications and with knowledge gaps in the underlying pathophysiology. Purpose To assess whether severe AS leads to prothrombotic left atrial (LA) flow changes with reduction of LA velocities and vorticity which do not normalise after AVR. Methods Seventy-seven participants without history of AF were recruited in this study: N=40 with severe AS (60% male; age 73 ±10 years, BMI 28 ±5 Kg/m2, median CHA2DS2VASc = 3) and N=37 controls with similar clinical risk profile (51% male; age 71 ±6 years, BMI 28 ±5 Kg/m2, median CHA2DS2VASc = 3). All participants underwent a cardiovascular magnetic resonance (CMR) scan to quantify left ventricular (LV) volumes, mass, and ejection fraction (LVEF), LV global longitudinal strain (GLS) as a sensitive marker of longitudinal dysfunction, LA maximum volume and emptying fraction (LAEF), LA blood flow peak velocity and vorticity by 4-dimensional flow CMR. Out of the 40 patients with severe AS, N=23 (58%) underwent a successful AVR and were followed-up with a repeat CMR scan at approximately 6 months after surgery. Results When compared to matched controls, patients with severe AS displayed significantly higher LV mass, smaller LA and LV volumes (i.e. concentric LV hypertrophy), significantly reduced GLS but higher (more hyperdynamic) LVEF (all p<0.05), as expected. LAEF (i.e. global LA function) was not significantly different from controls but despite this, severe AS was associated with slightly lower LA peak flow velocity (mean 0.25 vs. 0.28 m/sec respectively) and lower LA flow vorticity (mean 16.6 vs. 21.1 radians) compared to controls (both p<0.05). Successful AVR led to complete regression of LV hypertrophy (p<0.05) with LV mass that was no longer different from controls (p>0.05). Nevertheless, post-AVR patients displayed a persistent adverse morpho-functional phenotype characterised by persistently smaller LA and LV volumes, persistently hyperdynamic LV function with reduced GLS (all p<0.05), as well as adverse LA flow changes with significantly lower velocity (mean 0.24 vs 0.28 m/sec, p=0.003) and vorticity when compared to controls (17.1 vs 21.1 radians, p<0.001). Conclusions Patients with severe AS display adverse LA physiology and structure (prothrombotic LA flow characteristics in smaller LA size) and LV changes (smaller size with reduced longitudinal function and hyperdynamic function), which persist after successful AVR despite normalisation of the LV hypertrophy.Figure 1

Decreased Liver Kinase B1 Expression and Impaired Angiogenesis in a Murine Model of Bronchopulmonary Dysplasia.

Bronchopulmonary dysplasia (BPD) is characterized by impaired lung alveolar and vascular growth. We investigated the hypothesis that neonatal exposure to hyperoxia leads to persistent BPD phenotype caused by decreased expression of liver kinase B1 (LKB1), a key regulator of mitochondrial function. We exposed mouse pups from Postnatal Day (P)1 through P10 to 21% or 75% oxygen. Half of the pups in each group received metformin or saline intraperitoneally from P1 to P10. Pups were killed at P4 or P10 or recovered in 21% O2 until euthanasia at P21. Lung histology and morphometry, immunofluorescence, and immunoblots were performed to detect changes in lung structure and expression of LKB1; downstream targets AMPK, PGC-1α, and electron transport chain (ETC) complexes; and Notch ligands Jagged 1 and delta-like 4. LKB1 signaling and invitro angiogenesis were assessed in human pulmonary artery endothelial cells (exposed to 21% or 95% O2 for 36 hours. Levels of LKB1, phosphorylated AMPK, PGC-1α, and ETC complexes were decreased in lungs at P10 and P21 in hyperoxia. Metformin increased LKB1, phosphorylated AMPK, PGC-1α, and ETC complexes at P10 and P21 in pups exposed to hyperoxia. Radial alveolar count was decreased, and mean linear intercept increased in pups exposed to hyperoxia at P10 and P21; these were improved by metformin. Lung capillary density was decreased in hyperoxia at P10 and P21 and was increased by metformin. In vitro angiogenesis was decreased in human pulmonary artery endothelial cells by 95% O2 and was improved by metformin. Decreased LKB1 signaling may contribute to decreased alveolar and vascular growth in a mouse model of BPD.

Sensitisation to Lep d 2- Lepidoglyphus destructor allergy in asthma and rhinitis.

Background. Mites are responsible for allergic diseases, such as asthma and rhinitis, in nearly 1.5% of the world population. It is known that nowadays, besides House Dust Mites (HDM), Storage Mites (SM) are important sensitisers in urban non-occupational dwellings, predominantly in the Mediterranean area. The main objective of our study was to analyse the clinical relevance of the most prevalent SM, Lepidoglyphus destructor, by assessing the relationship between sensitisation to the major molecular allergen Lep d 2 and allergic respiratory disease phenotype in an urban population monosensitised to this molecular allergen. Methods. Cross-sectional study which included consecutive patients evaluated in our Allergy Department between 2012 and 2018, from urban non-occupational setting, with rhinitis and/or asthma, perennial symptoms and proven sensitisation to SM Lep d 2, tested using ImmunoCAP Immuno Solid-phase Allergen Chip (ISAC) technology, which detects 112 allergens. Results. A total of 37 allergic patients were included (23 female), aged 20.1 ± 12.8, min-max 5-58 years-old. The molecular sensitisation profile showed 18.9% of L. destructor mite monosensitised patients, having only sIgE to Lep d 2. This subset of patients mainly had allergic rhinitis, with 28.6% also being asthmatic. Regarding severity, most patients showed a persistent moderate phenotype of respiratory disease. The mean value of Lep d 2 sIgE was 8.3 ± 9.8 ISU-E, lower than in mite polysensitised patients (21.7 ± 21.5, p = 0.049). Conclusions. Our proof-of-concept study focused on Lep d 2 monosensitisation, showed that SM may have clinical relevance in perennial allergic asthma and rhinitis, and should be taken into account when assessing and treating allergic respiratory disease. Conclusions. The present survey demonstrated that Italian primary care pediatricians accomplish ARIA guidelines and adapt treatment on the basis of the intensity of symptoms. Corticosteroids and antihistamines are the most common prescribed medications. Nasal lavages are also popular.

Acute early life stress alters threat processing in adult rats.

Individuals diagnosed with stress-related psychiatric disorders in adulthood are likely to have experienced early life stress, suggesting that early adversity is an important vulnerability factor in the subsequent development of trauma- and anxiety-related psychiatric illness. It is important to develop animal models of psychiatric dysfunction to determine evident vulnerability considerations, potential biomarkers, and novel treatment avenues to improve the human condition. In our model of acute early life stress (aELS), 15 footshocks are delivered in a single session on postnatal day 17. The following experiments investigated the persistent impacts of our aELS procedure on stress-enhanced fear learning, anxiety-related behaviors, maintenance of fear, and resistance to extinction in adult male and female rats. The findings from these experiments demonstrate that our aELS procedure yields enhanced fear learning and increased anxiety. This enhanced fear is maintained over time, yet it extinguishes normally. Taken together, these results demonstrate that exposure to 15 footshocks during a single session early in life (postnatal day 17) recapitulates a number of important features of trauma- and anxiety-related disorder symptomatology, but not others. Future studies are needed to determine the persistent physiological phenotypes resulting from aELS and the neurobiological mechanisms that mediate these long-term changes. (PsycInfo Database Record (c) 2025 APA, all rights reserved).

Persistent Atrial Fibrillation Phenotypes and Ablation Outcomes: Persistent From Outset vs Progression From Paroxysmal AF

BackgroundMany patients with persistent atrial fibrillation (PsAF) have progressed from an initial paroxysmal phenotype; however, there are patients in whom atrial fibrillation (AF) is persistent at diagnosis. Relatively little is known about this subgroup, but prior observational studies have suggested these patients have worse outcomes with ablation. ObjectivesThis study sought to: 1) assess demographic and electrophysiologic characteristics of patients with PsAF at first diagnosis compared with those with who have progressed from paroxysmal atrial fibrillation (PAF); and 2) assess the impact of pattern of AF at diagnosis on recurrence post ablation. MethodsCAPLA (Catheter Ablation for persistent atrial fibrillation: A Multicentre randomised trial of Pulmonary vein isolation [PVI] vs PVI with posterior Left Atrial wall isolation [PWI]) was a multicenter trial that randomized patients with PsAF to PVI plus PWI or PVI alone. Follow-up was 12 months. Outcomes were assessed after a 3-month blanking period. ResultsA total of 334 patients were included (median age 65.6 years, 23.1% female), 194 (58.1%) had PsAF at first AF diagnosis and 140 (41.9%) had PAF. Patients with PsAF at diagnosis were younger (age 64.0 vs 67.7 years, P = 0.005), had higher rates of heart failure (P < 0.001), and lower left ventricular ejection fraction (54.5% IQR: 40-60 vs 60% IQR: 50-61, P = 0.007). AF recurrence occurred in 85 (43.8%) with PsAF at diagnosis and 70 (50%) with PAF at diagnosis. PsAF at diagnosis was not associated with risk of recurrence on univariable (HR: 0.802; 95% CI: 0.585-1.101; P = 0.173) or multivariable analysis (HR: 0.922; 95% CI: 0.647-1.312; P = 0.650). Median AF burden was 0% in both groups (P = 0.125). There was no difference in left atrial size (P = 0.337) or bipolar voltage (P = 0.579) between the groups. ConclusionsIn the CAPLA cohort of patients, pattern of AF at first diagnosis did not influence post-ablation rate of AF recurrence or AF burden. (Catheter Ablation for persistent atrial fibrillation: A Multicentre randomised trial of Pulmonary vein isolation [PVI] vs PVI with posterior Left Atrial wall isolation [PWI]; ACTRN12616001436460)

Metabolic specialization drives reduced pathogenicity in Pseudomonas aeruginosa isolates from cystic fibrosis patients.

Metabolism provides the foundation for all cellular functions. During persistent infections, in adapted pathogenic bacteria metabolism functions radically differently compared with more naïve strains. Whether this is simply a necessary accommodation to the persistence phenotype or if metabolism plays a direct role in achieving persistence in the host is still unclear. Here, we characterize a convergent shift in metabolic function(s) linked with the persistence phenotype during Pseudomonas aeruginosa colonization in the airways of people with cystic fibrosis. We show that clinically relevant mutations in the key metabolic enzyme, pyruvate dehydrogenase, lead to a host-specialized metabolism together with a lower virulence and immune response recruitment. These changes in infection phenotype are mediated by impaired type III secretion system activity and by secretion of the antioxidant metabolite, pyruvate, respectively. Our results show how metabolic adaptations directly impinge on persistence and pathogenicity in this organism.

Decoding Bacterial Persistence: Mechanisms and Strategies for Effective Eradication.

The ability of pathogenic bacteria to evade antibiotic treatment is an intricate and multifaceted phenomenon. Over the years, treatment failure among patients due to determinants of antimicrobial resistance (AMR) has been the focal point for the research and development of new therapeutic agents. However, the survival of bacteria by persisting under antibiotic stress has largely been overlooked. Bacterial persisters are a subpopulation of sensitive bacterial cells exhibiting a noninheritable drug-tolerant phenotype. They are linked to the recalcitrance of infections in healthcare settings, in turn giving rise to AMR variants. The importance of bacterial persistence in recurring infections has been firmly recognized. Fundamental work over the past decade has highlighted numerous unique tolerance factors contributing to the persister phenotype in many clinically relevant pathogens. This review summarizes contributing factors that could aid in developing new strategies against bacterial antibiotic persisters.

Adult Offspring of Deer Mouse Breeding Pairs Selected for Normal and Compulsive-Like Large Nesting Expression Invariably Show the Same Behavior Without Prior In-Breeding.

Obsessive-compulsive disorder is a neuropsychiatric condition with notable genetic involvement. Against this background, laboratory-housed deer mice of both sexes varyingly present with excessive and persistent large nesting behavior (LNB), which has been validated for its resemblance of clinical compulsivity. Although LNB differs from normal nesting behavior (NNB) on both a biological and cognitive level, it is unknown to what extent the expression of LNB and NNB is related to familial background. Here, we randomly selected 14 NNB- and 14 LNB-expressing mice (equally distributed between sexes) to constitute 7 breeding pairs of each phenotype. Pairs were allowed to breed two successive generations of offspring, which were raised until adulthood (12 weeks) and assessed for nesting expression. Remarkably, our findings show that offspring from LNB-expressing pairs build significantly larger nests compared to offspring from NNB-expressing pairs and the nesting expression of the offspring of each breeding pair, irrespective of parental phenotype or litter, is family specific. Collectively, the results of this investigation indicate that LNB can be explored for its potential to shed light on heritable neurocognitive mechanisms that may underlie the expression of specific persistent behavioral phenotypes.

Bacteriophage-driven emergence and expansion of Staphylococcus aureus in rodent populations.

Human activities such as agriculturalization and domestication have led to the emergence of many new pathogens via host-switching events between humans, domesticated and wild animals. Staphylococcus aureus is a multi-host opportunistic pathogen with a global healthcare and economic burden. Recently, it was discovered that laboratory and wild rodents can be colonised and infected with S. aureus, but the origins and zoonotic potential of rodent S. aureus is unknown. In order to trace their evolutionary history, we employed a dataset of 1249 S. aureus genome sequences including 393 of isolates from rodents and other small mammals (including newly determined sequences for 305 isolates from 7 countries). Among laboratory mouse populations, we identified multiple widespread rodent-specific S. aureus clones that likely originated in humans. Phylogeographic analysis of the most common murine lineage CC88 suggests that it emerged in the 1980s in laboratory mouse facilities most likely in North America, from where it spread to institutions around the world, via the distribution of mice for research. In contrast, wild rodents (mice, voles, squirrels) were colonized with a unique complement of S. aureus lineages that are widely disseminated across Europe. In order to investigate the molecular basis for S. aureus adaptation to rodent hosts, genome-wide association analysis was carried out revealing a unique complement of bacteriophages associated with a rodent host ecology. Of note, we identified novel prophages and pathogenicity islands in rodent-derived S. aureus that conferred the potential for coagulation of rodent plasma, a key phenotype of abscess formation and persistence. Our findings highlight the remarkable capacity of S. aureus to expand into new host populations, driven by the acquisition of genes promoting survival in new host-species.

A high-throughput assay identifies molecules with antimicrobial activity against persister cells.

Introduction. Persister cells are transiently non-growing antibiotic-tolerant bacteria that cause infection relapse, and there is no effective antibiotic therapy to tackle these infections.Gap statement. High-throughput assays in drug discovery are biased towards detecting drugs that inhibit bacterial growth rather than killing non-growing bacteria. A new and simple assay to discover such drugs is needed.Aim. This study aims to develop a simple and high-throughput assay to identify compounds with antimicrobial activity against persister cells and use it to identify molecular motifs with such activity.Methodology. We quantified Staphylococcus aureus persister cells by enumeration of colony forming units after 24 h ciprofloxacin treatment. We first quantified how the cell concentration, antibiotic concentration, growth phase and presence/absence of nutrients during antibiotic exposure affected the fraction of persister cells in a population. After optimizing these parameters, we screened the antimicrobial activity of compound fragments to identify molecular structures that have activity against persister cells.Results. Exponential- and stationary-phase cultures transferred to nutrient-rich media displayed a bi-phasic time-kill curve and contained 0.001-0.07% persister cells. A short rifampicin treatment resulted in 100% persister cells for 7 h, after which cells resumed activity and became susceptible. Stationary-phase cultures displayed a low but constant death rate but ultimately resulted in similarly low survival rates as the exponential-phase cultures after 24 h ciprofloxacin treatment. The persister phenotype was only maintained in most of the population for 24 h if cells were transferred to a carbon-free minimal medium before exposure to ciprofloxacin. Keeping cells starved enabled the generation of high concentrations of S. aureus cells that tolerate 50× MIC ciprofloxacin, and we used this protocol for rapid screening for biocidal antibiotics. We identified seven compounds from four structural clusters with activity against antibiotic-tolerant S. aureus. Two compounds were moderately cytotoxic, and the rest were highly cytotoxic.Conclusion. Transferring a stationary-phase culture to a carbon-free minimal medium for antimicrobial testing is a simple strategy for high-throughput screening for new antibiotics that kill persister cells. We identified molecule fragments with such activity, but further screening is needed to identify motifs with lower general cytotoxicity.

Chronic ecologically relevant stress effects on reverse-translated touchscreen assays of reward responsivity and attentional processes in male rats: Implications for depression.

Stagnation in the development of novel therapeutic strategies for treatment-resistant depression has encouraged continued interest in improving preclinical methods. One tactic prioritizes the reverse translation of behavioral tasks developed to objectively quantify depressive phenotypes in patient populations for their use in laboratory animals via touchscreen technology. After cross-species concordance in task outcomes under healthy conditions is confirmed, construct validity can be further enhanced by identifying environmental stressors that reliably produce deficits in task performance that resemble those in depressive participants. The present studies characterized in male rats the ability of two chronic ecologically relevant stressors, inescapable ice water or isolated restraint, to produce depressive-like behavioral phenotypes in the Probabilistic Reward Task (PRT) and Psychomotor Vigilance Task (PVT). These tasks previously have been reverse-translated using touchscreen technology for rodents and nonhuman primates to objectively quantify, respectively, reward responsivity (anhedonia) and attentional processes (impaired cognitive function), each of which are core features of major depressive disorder. In the PRT, both inescapable ice water and isolated restraint produced persistent anhedonic phenotypes compared to non-stressed control performance (i.e., significantly blunted response bias for the richly rewarded stimulus). In the PVT, both chronic stressors impaired attentional processing, revealed by increases in titrated reaction times; however, these deficits largely subsided by the end of the chronic condition. Taken together, these findings confirm the ability of reverse-translated touchscreen tasks to effectively generate behavioral phenotypes that exhibit expected deficits in performance outcomes following exposure to chronic ecologically relevant stress. In turn, this approach is well positioned to appraise the ability of candidate therapeutics to attenuate or reverse such behavioral deficits and, thereby, contribute to preclinical medications development for treatment-resistant depression.

Tracking Chlamydia – Host interactions and antichlamydial activity in Caenorhabditis elegans

The fading efficacy of antibiotics is a growing global health concern due to its life-threatening consequences and increased healthcare costs. Non-genetic mechanisms of antimicrobial resistance, such as those employed by Chlamydia pneumoniae and Chlamydia trachomatis, complicate treatment as these bacteria can enter a non-replicative, persistent state under stress, evading antibiotics and linking to inflammatory conditions. Understanding chlamydial persistence at the molecular level is challenging, and new models for studying Chlamydia-host interactions in vivo are urgently needed. Caenorhabditis elegans offers an alternative given its immune system and numerous orthologues of human genes. This study established C. elegans as an in vivo model for chlamydial infection. Both Chlamydia species reduced the worm's lifespan, their DNA being detectable at three- and six-days post-infection. Azithromycin at its MIC (25 nM) failed to prevent the infection-induced lifespan reduction, indicating a persister phenotype. In contrast, the methanolic extract of Schisandra chinensis berries showed anti-chlamydial activity both in vitro (in THP-1 macrophages) and in vivo, significantly extending the lifespan of infected C. elegans and reducing the bacterial load. Moreover, S. chinensis increased the transcriptional activity of SKN-1 in the worms, but was unable to impact the bacterial load or lifespan in a sek-1 defective C. elegans strain. In summary, this study validated C. elegans as a chlamydial infection model and showcased S. chinensis berries' in vivo anti-chlamydial potential, possibly through SEK/SKN-1 signaling modulation.

A Novel Murine Model for Lupus-Like Ocular Chronic Graft-Versus-Host Disease.

Lupus-like chronic graft-versus-host disease (cGVHD) has been previously described, but the ocular findings have not been elucidated. Recipient mice in a lupus-like cGVHD model manifested notable and persistent ocular surface phenotypes. Herein, we further explored immunopathogenic mechanisms underlying these ocular phenotypes. A previously described lupus-like cGVHD model was established by intraperitoneal injection of splenocytes from bm12 mice into C57BL/6J mice. Systemic findings were evaluated for the presence of splenomegaly, proteinuria, and autoantibodies. Comprehensive evaluations were conducted on ocular manifestations and immunopathological features in this model. The lupus-like cGVHD model was successfully constructed 2 weeks post-transplantation. The recipient mice developed lupus-like phenotypes, including splenomegaly, proteinuria, and increased autoantibodies, and their ocular presentations included corneal epithelial defects and decreased tear secretion. Histological analysis revealed a reduction in corneal nerve fiber density and corneal endothelial cells, along with conjunctival fibrosis and loss of goblet cells. Moreover, cGVHD induced progressive aggravation of immune cell infiltration and fibrosis in the lacrimal glands. RNA-Sequencing (RNA-seq) results of the lacrimal glands demonstrated that the differentially expressed genes (DEGs) between the control and cGVHD groups were associated with GVHD pathways. Immune infiltration analysis using RNA-seq and flow cytometry confirmed that CD8+ T lymphocytes predominantly constituted the inflammatory infiltrating cells within the lacrimal glands. This lupus-like cGVHD model (bm12→C57BL/6J) exhibited persistent ocular surface manifestations, characterized by immune infiltration of CD8+ T lymphocytes in the lacrimal glands. Thus, this ocular cGVHD model may be used to explore the underlying mechanisms and discover novel therapeutic interventions.