Phage therapy combined with Gum Karaya injectable hydrogels for treatment of methicillin-resistant Staphylococcus aureus deep wound infection in a porcine model.

Skin and soft tissue infections (SSTIs) represent a significant healthcare challenge, particularly in the context of increasing antibiotic resistance. This study investigates the efficacy of a novel therapeutic approach combining bacteriophage (phage) therapy with a gum Karaya (GK)-based hydrogel delivery system in a porcine model of deep staphylococcal SSTIs. The study exploits the lytic activity and safety of the Staphylococcus phage 812K1/420 of the Kayvirus genus, which is active against methicillin-resistant Staphylococcus aureus (MRSA). The GK injectable hydrogels and hydrogel films, developed by our research group, serve as effective, non-toxic, and easy-to-apply delivery systems, supporting moist wound healing and re-epithelialization. In the porcine model, the combined treatment showed asynergistic effect, leading to a significant reduction in bacterial load (2.5 log CFU/gram of tissue) within one week. Local signs of inflammation were significantly reduced by day 8, with clear evidence of re-epithelialization and wound contraction. Importantly, no adverse effects of the GK-based delivery system were observed throughout the study. The results highlight the potential of this innovative therapeutic approach to effectively treat deep staphylococcal SSTIs, providing a promising avenue for further research and clinical application in the field of infections caused by antibiotic-resistant bacteria.

Large-scale mutational analysis identifies UNC93B1 variants that drive TLR-mediated autoimmunity in mice and humans.

Nucleic acid-sensing Toll-like receptors (TLR) 3, 7/8, and 9 are key innate immune sensors whose activities must be tightly regulated to prevent systemic autoimmune or autoinflammatory disease or virus-associated immunopathology. Here, we report a systematic scanning-alanine mutagenesis screen of all cytosolic and luminal residues of the TLR chaperone protein UNC93B1, which identified both negative and positive regulatory regions affecting TLR3, TLR7, and TLR9 responses. We subsequently identified two families harboring heterozygous coding mutations in UNC93B1, UNC93B1+/T93I and UNC93B1+/R336C, both in key negative regulatory regions identified in our screen. These patients presented with cutaneous tumid lupus and juvenile idiopathic arthritis plus neuroinflammatory disease, respectively. Disruption of UNC93B1-mediated regulation by these mutations led to enhanced TLR7/8 responses, and both variants resulted in systemic autoimmune or inflammatory disease when introduced into mice via genome editing. Altogether, our results implicate the UNC93B1-TLR7/8 axis in human monogenic autoimmune diseases and provide a functional resource to assess the impact of yet-to-be-reported UNC93B1 mutations.

Increasing prevalence of celiac disease - where to look for answers?

Celiac disease or gluten-sensitive enteropathy is a relevant health concern in today's world. Three prerequisites need to be met to trigger the disease, namely a genetic predisposition, gluten consumption, and environmental factors. Retrospective studies conducted across all age groups have ruled out the possibility that improved diagnostic methods were behind the increased prevalence. Since the genetic predisposition is more or less constant in the population, it is assumed that external factors may play a major role in this increase. Although it is generally believed that modern wheat varieties are to be blamed for the increase in gluten intolerance-related diseases, this assumption is refuted based on the analysis of the current and 100-year-old varieties. However, the increased prevalence could be related to modern lifestyles, changes in food preparation technology or composition, disruption of the intestinal barrier in viral disease, and other factors leading to intestinal dysbiosis. A possible preventive strategy in predisposed individuals could be the avoidance of gluten from the diet when ill, especially with a viral infection. This article openup a new perspective on the currently common autoimmune disease.

Insurance denials and diagnostic rates in a pediatric genomic research cohort.

PURPOSE: This study aimed to assess the amount and types of clinical genetic testing denied by insurance and the rate of diagnostic and candidate genetic findings identified through research in patients who faced insurance denials. METHODS: Analysis consisted of review of insurance denials in 801 patients enrolled in a pediatric genomic research repository with either no previous genetic testing or previous negative genetic testing result identified through cross-referencing with insurance prior-authorizations in patient medical records. Patients and denials were also categorized by type of insurance coverage. Diagnostic findings and candidate genetic findings in these groups were determined through review of our internal variant database and patient charts. RESULTS: Of the 801 patients analyzed, 147 had insurance prior-authorization denials on record (18.3%). Exome sequencing and microarray were the most frequently denied genetic tests. Private insurance was significantly more likely to deny testing than public insurance (odds ratio = 2.03 [95% CI = 1.38-2.99] P = .0003). Of the 147 patients with insurance denials, 53.7% had at least 1 diagnostic or candidate finding and 10.9% specifically had a clinically diagnostic finding. Fifty percent of patients with clinically diagnostic results had immediate medical management changes (5.4% of all patients experiencing denials). CONCLUSION: Many patients face a major barrier to genetic testing in the form of lack of insurance coverage. A number of these patients have clinically diagnostic findings with medical management implications that would not have been identified without access to research testing. These findings support re-evaluation of insurance carriers' coverage policies.

Neurocognitive correlates of the varied domains of outcomes at 20 year follow-up of first-episode psychosis.

Little is known about long-term outcomes of the first episode of psychosis (FEP) other than in the symptomatic domain. We hypothesised that cognitive impairment is associated with poorer multi-domain outcomes at a long-term follow-up of FEP patients. We followed-up 172 FEP patients for a mean of 20.3 years. Ten outcome dimensions were assessed (symptomatic, functional and personal recovery, social disadvantage, physical health, suicide attempts, number of episodes, current drug use, chlorpromazine equivalent doses (CPZ), and schizophrenia/schizoaffective disorder final diagnosis). Cognition was assessed at follow-up. Processing speed and verbal memory deficits showed significant associations with poor outcomes on symptomatic, social functioning, social disadvantage, higher number of episodes, and higher CPZ. Significant associations were found between visual memory impairments were significantly associated with low symptomatic and functional recovery, between attentional deficits and a final diagnosis of schizophrenia/schizoaffective disorder, and between social cognition deficits and poor personal recovery.Lower cognitive global scores were significantly associated with all outcome dimensions except for drug abuse and physical status. Using multiple outcome dimensions allowed for the inclusion of the patients' perspective and other commonly neglected outcome measures. Taken together, cognitive impairment in FEP patients is strongly related to poor performance on several outcome dimensions beyond symptomatic remission.

Genomic answers for children: Dynamic analyses of >1000 pediatric rare disease genomes.

PURPOSE: This study aimed to provide comprehensive diagnostic and candidate analyses in a pediatric rare disease cohort through the Genomic Answers for Kids program. METHODS: Extensive analyses of 960 families with suspected genetic disorders included short-read exome sequencing and short-read genome sequencing (srGS); PacBio HiFi long-read genome sequencing (HiFi-GS); variant calling for single nucleotide variants (SNV), structural variant (SV), and repeat variants; and machine-learning variant prioritization. Structured phenotypes, prioritized variants, and pedigrees were stored in PhenoTips database, with data sharing through controlled access the database of Genotypes and Phenotypes. RESULTS: Diagnostic rates ranged from 11% in patients with prior negative genetic testing to 34.5% in naive patients. Incorporating SVs from genome sequencing added up to 13% of new diagnoses in previously unsolved cases. HiFi-GS yielded increased discovery rate with >4-fold more rare coding SVs compared with srGS. Variants and genes of unknown significance remain the most common finding (58% of nondiagnostic cases). CONCLUSION: Computational prioritization is efficient for diagnostic SNVs. Thorough identification of non-SNVs remains challenging and is partly mitigated using HiFi-GS sequencing. Importantly, community research is supported by sharing real-time data to accelerate gene validation and by providing HiFi variant (SNV/SV) resources from >1000 human alleles to facilitate implementation of new sequencing platforms for rare disease diagnoses.

Enzyme-based treatment of skin and soft tissue infections.

Inflammatory diseases of the skin and soft tissues are an important group of human infections. The most common causes are the bacteria Staphylococcus aureus and Streptococcus pyogenes. Given the growing resistance of these pathogens to antimicrobials, the current research focuses on the search for novel therapeutic options that would be effective against infections refractory to conventional antimicrobials. A promising alternative is the use of enzyme-based antimicrobials (enzybiotics) that degrade the bacterial cell wall. They target the specific pathogen but do not affect the skin microbiome, thus helping the healing process. As enzymes can be poorly soluble, unstable, or subject to rapid elimination from the body, efforts are made to create biobetters, i.e., enzymes with improved characteristics. Emphasis is also put on the development of novel enzybiotic carriers or wound healing dressings with integrated enzymes.

De novo heterozygous missense and loss-of-function variants in CDC42BPB are associated with a neurodevelopmental phenotype.

CDC42BPB encodes MRCKß (myotonic dystrophy-related Cdc42-binding kinase beta), a serine/threonine protein kinase, and a downstream effector of CDC42, which has recently been associated with Takenouchi-Kosaki syndrome, an autosomal dominant neurodevelopmental disorder. We identified 12 heterozygous predicted deleterious variants in CDC42BPB (9 missense, 2 frameshift, and 1 nonsense) in 14 unrelated individuals (confirmed de novo in 11/14) with neurodevelopmental disorders including developmental delay/intellectual disability, autism, hypotonia, and structural brain abnormalities including cerebellar vermis hypoplasia and agenesis/hypoplasia of the corpus callosum. The frameshift and nonsense variants in CDC42BPB are expected to be gene-disrupting and lead to haploinsufficiency via nonsense-mediated decay. All missense variants are located in highly conserved and functionally important protein domains/regions: 3 are found in the protein kinase domain, 2 are in the citron homology domain, and 4 in a 20-amino acid sequence between 2 coiled-coil regions, 2 of which are recurrent. Future studies will help to delineate the natural history and to elucidate the underlying biological mechanisms of the missense variants leading to the neurodevelopmental and behavioral phenotypes.

Plectin repeats and modules: strategic cysteines and their presumed impact on cytolinker functions.

Plectin, a member of the cytolinkers protein family, plays a crucial role in cells as a stabilizing element of cells against mechanical stress. Its absence results in muscular dystrophy, skin blistering, and signs of neuropathy. The C-terminal domain of plectin contains several highly homologous repeat domains that also occur in other cytolinkers. Secondary structure analysis revealed that the building block of these domains, the PLEC repeat, is similar to the ankyrin repeat. We present a model that attempts to explain how the C-terminal domain, which comprises approximately 1900 amino acid, could be stabilized to maintain its structural integrity even under extensive mechanical stress. In this model, larger solenoid modules formed from PLEC repeats can be disulfide-bridged via conserved cysteines. Our hypothesis suggests that this process could be mediated by cytoplasmic NOS-generated products, such as the radical peroxynitrite. Reinforcement of molecular structure would provide a rationale why during exercising or physical stress radicals are formed without necessarily being deleterious. This article contains supplementary material that may be viewed at the BioEssays website at http://www.interscience.wiley.com/jpages/0265-9247/suppmat/23/v23_11.1064.html.

IVR Test & Survey: a computer program to collect data via computerized telephonic applications.

Several studies have demonstrated the advantages of using interactive voice response (IVR) technology to collect self-report data from research participants and recipients of psychological/medical services. IVR allows participants to phone a computer and respond to recorded questions by pressing the appropriate touch-tone keys on their telephone. Because this technology offers substantial benefits in terms of cost and efficiency, it is surprising that it has not been more widely utilized by researchers and practitioners. Along with the automation of the administration and scoring of tests or surveys, IVR provides for questioning to be adapted to the participants' responses. One possible explanation for the failure to exploit this technology is the absence of easy-to-use software that does not require programming knowledge. This article describes IVR Test & Survey, a program to facilitate the administration, scoring, and analysis of information collected with the use of IVR technology.

Molecular characterization of the Thermomonospora curvata aglA gene encoding a thermotolerant alpha-1,4-glucosidase.

The cloning, sequencing and structural characterization of a gene encoding a thermostable alpha-1,4-glucosidase from Thermomonospora curvata is described. DNA sequence analysis revealed four open reading frames designated aglA, aglR, aglE and aglF. The aglA gene encodes a thermostable alpha-1,4-glucosidase from T. curvata and is situated between two genes, aglR and aglE. Genes aglA, aglE and aglF are transcribed in the same direction, while aglR is transcribed in the opposite direction. By comparing the amino acid sequence of the alpha-1,4-glucosidase from T. curvata with other alpha-glucanases, it appears that the enzyme is a member of the alpha-amylase family. The proteins of this family have an (alpha/beta)8 barrel super secondary structure. The topology of the alpha-1,4-glucosidase was predicted by computer-assisted analysis. The topology of the secondary structures of the alpha-1,4-glucosidase resembles the structure of barley alpha-amylase, but the primary structure resembles most closely the oligo-1,6-glucosidase from Bacillus cereus. Putative catalytic residues (D221, E281 and D343) and calcium binding residues (N116, E179, D191, H224 or G225) are proposed.

Genotypic and phenotypic similarity of multiresistant Acinetobacter baumannii isolates in the Czech Republic.

The diversity of 103 clinical isolates of the Acinetobacter calcoaceticus-Acinetobacter baumannii complex obtained between 1991 and 1997 from 17 Czech hospitals was studied by ribotyping, biotyping, plasmid profiling and antibiotic susceptibility testing. According to the EcoRI ribotypes, all but one of these isolates were identified to the DNA group level: 77 isolates were allocated to DNA group 2 (A. baumannii), 14 to DNA group 3, 10 to DNA group 13 sensu Tjernberg and Ursing and one to DNA group 1 (A. calcoaceticus). In total, 50 different EcoRI ribotypes and 10 biotypes were observed. Plasmids were found in 92% of the isolates and a high variability in plasmid profiles was found in isolates of the same DNA group. The combination of typing profiles allowed two predominant groups (termed A and B) to be distinguished among the A. baumannii isolates (37 and eight isolates, respectively) that shared a specific ribotype and were highly similar in other properties. These two groups comprised both sporadic and outbreak isolates and were found in most localities. Group A and B isolates were markedly more resistant to antibiotics than most of the remaining isolates, thus representing 85% of all multiresistant isolates. The features of groups A and B corresponded to those of two epidemic clones identified recently among hospital strains in north-western Europe.

Neuropsychological performance of adults with attention deficit hyperactivity disorder (ADHD): diagnostic classification estimates for measures of frontal lobe/executive functioning.

ADHD adults (N = 26) were compared to normal controls (N = 26) on 6 neuropsychological measures believed sensitive to frontal lobe-executive functioning. MANOVA analyses and subsequent univariate tests indicated that most of the neuropsychological measures discriminated between the two groups. To address clinical significance diagnostic classification rates were also generated for each measure individually, and for the battery as a whole. Levels of positive predictive power (PPP) for each of the 6 measures (83-100%) indicated that abnormal scores on these tests were good predictors of ADHD. However, estimates of negative predictive power (NPP) suggested that normal scores poorly predicted the absence of ADHD. When classification rates were calculated for the overall battery classification accuracy improved substantially. Thus, neuropsychological tests can differentiate adults suffering from ADHD from adults without ADHD, while also providing good classification accuracy. Finally, the pattern of neurobehavioral impairments exemplified through the Summary Index scores was interpreted as consistent with conceptualizations of ADHD depicting mild neurologic dysfunction in networks associated with the frontal lobes.

Kytococcus sedentarius (formerly Micrococcus sedentarius) and Dermacoccus nishinomiyaensis (formerly Micrococcus nishinomiyaensis) produce monensins, typical Streptomyces cinnamonensis metabolites.

The environmental isolate Kytococcus sedentarius TR-2 was found to be a new producer of the oligoketide antibiotics monensin A and B. Electron microscopic studies demonstrated that the TR-2 strain had coccoid cells and DNA analysis revealed no close relationship to Streptomyces cinnamonensis, a typical monensin producer. Production of monensins was also proven with six culture collection K. sedentarius strains and three Dermacoccus nishinomiyaensis strains. The secondary metabolism of micrococci demonstrates a high degree of instability. Biosynthesis of monensins by micrococci endorses a phylogenetic relationship to Streptomyces spp.

A deduced Thermomonospora curvata protein containing serine/threonine protein kinase and WD-repeat domains.

The gene pkwA coding for a typical WD-repeat protein was found in the chromosome of the bacterium Thermomonospora curvata CCM 3352. Until now WD-repeat proteins were through to be confined to eukaryotes.

The interaction of substituted 2-phenylquinoline intercalators with poly(A).poly(U): classical and threading intercalation modes with RNA.

The interaction of a series of 2-phenylquinoline derivatives with RNA was investigated by means of viscometric, pKa, spectroscopic, binding, Tm, and kinetic methods. Compounds 1, 2, and 3 have a piperazyl substituent at the para, meta, or ortho position, respectively, while 4 has an unsubstituted phenyl ring. The pKa results suggest that 1 has three charges, 2 and 3 have more than two charges, and 4 has two charges at pH 6.2. Spectroscopic and Tm results indicate that 1 binds more strongly to RNA than 2-4. Kinetic and modeling results indicate that 1 is a threading intercalator while 2 and 4 are classical intercalators. All experimental results indicate that 3, which has a large twist between the phenyl and quinoline rings, binds weakly with RNA.

The relationship between adult sexual adjustment and childhood experiences regarding exposure to nudity, sleeping in the parental bed, and parental attitudes toward sexuality.

The relationship between adult sexual functioning and childhood experiences with exposure to nudity, sleeping in the parents' bed, and parental attitudes toward sexuality was examined. Although a variety of experts have provided their opinion on this issue, empirical research on this topic has been lacking. In this study, male and female college students were asked to retrospectively report on the frequency of sleeping in the parental bed as a child, the frequency of seeing others nude during childhood, and parental attitudes regarding sexuality. Information on current sexual functioning and adjustment was also obtained. The results suggest that childhood experiences with exposure to nudity and sleeping in the parental bed are not adversely related to adult sexual functioning and adjustment. In fact, there is modest support that these childhood experiences are positively related to indices of adjustment. Results also suggest that a positive attitude toward sexuality can be beneficial for a child's comfort with his/her sexuality. Finally, examination of gender differences revealed that male and female experience paternal attitudes toward sexuality differently but are similar in their perceptions of maternal attitudes.