Clinical and immunological assessment of APDS2 with features of the SHORT syndrome related to a novel mutation in PIK3R1 with reduced penetrance

Monoallelic loss-of-function (LOF) mutations in the phosphatidylinositol 3-kinase (PIK3R1) gene affecting the inter-Src homology 2 domain of the p85α regulatory subunit of phosphoinositide-3-kinase δ (PI3Kδ) cause the activated PI3K δ syndrome (APDS2). APDS2 is defined as a primary antibody deficiency, developmental abnormalities within the B and T lymph cell compartments, and immune dysregulation. The genetic defect of APDS2 is shared with that of the SHORT syndrome, characterized by short stature, joint hyperextensibility, ocular depression, Rieger anomaly, and delayed tooth eruption. LOF variants in an intronic splice site (c.1425+1G.C/A/T) in the PI3KR1 gene have been identified in patients affected with both APDS2 and SHORT syndrome. Herein, we report a novel c.1644-1648del (p.Asp548Glufs*6) variant in a pediatric patient with the APDS2-related immunodeficiency, who presents with mild phenotypic fea-tures of the SHORT syndrome, congenital chest wall deformity, and IgE-mediated food allergy. The same variant was also identified in the patient’s hitherto asymptomatic mother, impli-cating an incomplete penetrance. Regular monitoring by a multidisciplinary team under the pediatric clinical immunologist’s supervision to implement appropriate diagnostic procedures and treatment modalities is of paramount importance. Further studies are required to better define the genotype-phenotype correlation in patients with the PIK3R1 gene mutations and to better delineate the mutual relationship between APDS2 and the SHORT syndrome (AU)

Distribution of classical enterotoxin genes in staphylococci from milk of cows with- and without mastitis and the cowshed environment.

The aim of this study was to analyze by PCR 185 isolates of Staphylococcus from milk of cows with- and without mastitis and from the cowsheds environment for their potential ability to produce five classical staphylococcal enterotoxins. Among S. aureus isolates 8 (32%) carried enterotoxin genes and only 2 of them had more than one gene. The enterotoxin genes were detected in 22 (13.7%) coagulase-negative staphylococci (CNS) isolates, among them in 9 (11.4%) isolates of S. xylosus, 5 (16.7%) S. sciuri, 3 (10.3%) S. epidermidis and in 5 (22.7%) Staphylococcus spp. In some CNS 2 or 3 genes were detected simultaneously. Among the investigated enterotoxin genes, sec was the most prevalent (70%). The genes encoding enterotoxin B and D were detected in 5 (16.7%) and 6 (20%) isolates, respectively. The lowest number of isolates had sea and see genes. The genes encoding enterotoxins were often identified in staphylococci from milk of cows with mastitis (73.4% of detected genes), while only 6 (20%) isolates from milk of cows without mastitis and 2 (6.6%) isolates from cowshed environment were positive for enterotoxin genes. The results showed that CNS from bovine milk, like S. aureus, carried enterotoxin genes and may pose a risk for public health.

Motility activity, slime production, biofilm formation and genetic typing by ERIC-PCR for Pseudomonas aeruginosa strains isolated from bovine and other sources (human and environment).

The molecular-typing strategy, ERIC-PCR was used in an attempt to determine the genomic relationship of 28 P. aeruginosa strains isolated from faeces of healthy bovine, bovine mastitis and from faeces of hospital patients as well as from environment. ERIC-PCR fingerprinting revealed large molecular differentiation within this group of isolates. Twenty two out of 28 strains tested generated unique patterns of DNA bands and only three genotypes consisted of two isolates each were identified. We also tested the P. aeruginosa isolates for their ability to form a biofilm on abiotic surfaces including polyvinylchloride and polystyrene. Different biofilm-forming abilities were demonstrated among strains; however, most of them (64.3%) showed moderate-biofilm forming ability. The strains with increased swimming and twitching motility displayed elevated biofilm formation. However, a negative correlation was found between slime and initial biofilm production. On the basis of the results obtained, we suggest that there are no major differences in phenotypic properties between P. aeruginosa strains isolated from different sources.

Virulence factors and ability of staphylococci from bovine milk and the cowshed environment to biofilm formation.

The aim of this study was to examine virulence factors and the ability of S. aureus and CNS species isolated from milk of cows with mastitis to form biofilm, and to compare them with virulence factors of staphylococci from milk of cows without mastitis and cowshed environment. Most of S. aureus strains from cows with mastitis showed haemolytic activity (93.9%), among them 72.7% and 21.2% produced alpha- and beta-haemolysin, respectively. S. aureus from cows with mastitis symptoms produced proteases (above 48%) and esterase (42.4%). The highly significant relationship between the number of S. xylosus strains producing haemolysins (62%) and the origin of these strains from milk of cows with mastitis was observed. The ability to produce proteases was significantly associated with S. sciuri from milk of cows with mastitis. The ability of biofilm formation by staphylococcal strains from milk of cows with mastitis was greater than in strains from milk of cows without mastitis and the difference was significant (p < or = 0.05). The highest percentage of strains from milk of cows with mastitis were weak biofilm formers (48.6%), while 40% and 11.4% of strains were moderate and strong biofilm producers, respectively. S. xylosus showed the highest ability to form biofilm, while the lowest ability to form biofilm was observed in S. aureus and S. epidermidis. In conclusion, production of exotoxins and enzymes, and ability of biofilm formation shown by many CNS isolated from milk of cows with mastitis symptoms indicates that these features are important in pathogenesis of this disease.

Common transcriptional effects in the mouse striatum following chronic treatment with heroin and methamphetamine.

The molecular alterations that underlie the long-lasting behavioural effects of drugs of abuse, such as psychomotor sensitization and physical dependence, are still not known. Moreover, it is not known which molecular effects are similar for addictive drugs from various pharmacological classes. In this study, we utilized whole-genome microarray profiling to evaluate the detailed time-course of transcriptional alterations in the mouse striatum during chronic treatment with heroin (HER) and methamphetamine (METH) and after period of spontaneous withdrawal. We identified 27 genes regulated by chronic drug administration. The overlap between lists of HER- and METH-induced genes was highly significant. The most substantial impact on the gene expression profile was observed for circadian genes (Per1, Per2 and Nr1d1). However, changing the treatment scheme from diurnal to nocturnal was sufficient to attenuate the drug-induced changes in circadian gene mRNA levels. Both of the drugs caused a dose-dependent induction in glucocorticoid-dependent genes with relatively long mRNA half-lives (Fkbp5, Sult1a1 and Plin4). The analysis also showed a drug-regulated group of transcripts enriched in the nucleus accumbens and includes well known (Pdyn, Cartpt and Rgs2) as well as new (Fam40b and Inmt) candidate genes. All identified alterations in the striatal transcriptome were transient and persisted up to 6 days after withdrawal. Behavioural sensitization, however, was maintained throughout the 12-day withdrawal period for both HER and METH. We suggest that transient gene expression alterations during drug treatment and in the early period of withdrawal are involved in the establishment of persistent neuroplastic alterations responsible for the development of drug addiction.

Phenotypic and genotypic antimicrobial resistance of staphylococci from bovine milk.

The aim of this study was to examine phenotypic and genotypic antimicrobial resistance of staphylococci from milk samples from cows with subclinical and clinical mastitis and from cows without mastitis symptoms to methicillin, tetracyclines, macrolides and lincosamides (ML). Of 207 strains, including 34 S. aureus and 173 coagulase-negative staphylococci (CNS), 11 (6.4%) CNS strains were phenotypically resistant to methicillin. The mecA gene was detected by PCR only in two S. xylosus strains and one strain of S. epidermidis and S. simulans. No methicillin-resistant S. aureus strains were observed. In methicillin-resistant strains with mecA, gene resistance to other investigated antibiotics was not observed. Phenotypic resistance to tetracycline was detected in 11.0% of CNS strains and 47.4% of them carried the tetK gene. Of 173 CNS strains studied, 27 (15.6%) were resistant to at least one ML antibiotic. The resistance gene ermC was detected in 55.5% of the 27 ML-resistant strains. The ermA and ermB genes were detected in 14.8% and 11.1% of ML-resistant CNS strains, respectively. Antimicrobial resistance to methicillin, tetracyclines and macrolides was detected more frequently in staphylococcal strains from clinical mastitis compared to animals with subclinical symptoms and without mastitis, while the resistance to lincosamides showed a similar frequency in all groups of cows. In conclusion, CNS species from bovine milk differ in phenotypic and genotypic antimicrobial resistance profiles, and the use of PCR technique alone for the detection of methicillin, macrolide, lincosamide and tetyracycline resistance in CNS from cattle is not reliable.

Antimicrobial resistance and genotypes of staphylococci from bovine milk and the cowshed environment.

Investigation of antimicrobial resistance and genetic relatedness of staphylococci from milk of cows with mastitis and cowshed environment was the aim of this study. Antimicrobial resistance against 14 antimicrobials were determined by using a disc diffusion method. Genetic similarity between the most frequently isolated species was analysed by PFGE (pulsed-field gel electrophoresis). Haemolytic activity, DNase, protease and esterase production was also investigated. Coagulase-negative Staphylococcus species were isolated from 30.8% of milk samples from cows with mastitis. The most frequently isolated species was Staphylococcus xylosus and yield of these organisms was significantly associated with milk of mastitis cows. S. epidermidis was a predominant penicillin-resistant species. High frequency of resistance to lincomycin was observed among isolates of S. sciuri (54.2%) and S. xylosus (25.9%) from cows with mastitis. PFGE (pulsed-field gel electrophoresis) analysis of 29 Staphylococcus aureus isolates showed the presence of 17 PFGE pulsotypes. Isolates of S. sciuri (n = 36) had unique PFGE patterns. Some S. xylosus isolates from milk and milker's hands had the same PFGE pulsotypes, and this observation could indicate that dairyman may be a potential source of the infection. The pulsotype of each of the remaining isolates of S. xylosus suggested that they might have come from common environmental sources; however, these isolates differed in antibiotic resistance pattern or virulence traits. Therefore, knowledge about antibiotic sensitivity pattern and virulence factors of a CNS isolate, besides its genotype, may be informative in tracking the source of the infection.

Analysis of occurrence of virulence genes among Yersinia enterocolitica isolates belonging to different biotypes and serotypes.

The 150 Y enterocolitica strains isolated from humans and from pigs belonged to biotypes 4 (68.7%), 1A (18.7%) and 2 (4%), or were biochemically untypeable (8.6%). Biotype 4 was comprised of Y. enterocolitica strains representing serotype O:3, within biotype 1A the strains either belonged to serotypes O:5 and O:6 or were untypeable, and biotype 2 was represented by the strains of serotype O:9. The strains which were biochemically untypeable belonged to serotypes O:5, O:6 and O:3. Among the strains tested there also were those of an unidentified biotype and serotype. Nearly all the strains of biotype 1A represented genotype ystB+myfA+, and few belonged to genotype ystB+. The presence of the ystB gene in the strains of biotype 1A and only occasional occurrence of the gene in the other biotypes makes ystB a distinguishing marker of biotype 1A. The strains of genotype ystA+ail+myfA+yadA+ predominated in biotype 4 (serotype O:3). The strains of biotype 2 (serotype O:9) represented genotype ystA+ail+myfA+, and the plasmid yadA gene was detected in some of them. Within the group of biochemically untypeable strains ystB- and myfA-specific PCR products were mainly obtained. The genotypes determined for the tested biotypes and serotypes of Y. enterocolitica, based upon the selected genes of virulence, can be applied as distinguishing markers and indicators of the potential virulence of Y. enterocolitica strains, excluding bioserotyping.

Natriuretic peptides in septic patients.

The natriuretic peptide family is comprised of atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), C-type natriuretic peptide (CNP), dendroaspis natriuretic peptide - DNP and urodilatin. They play a role in the diagnosis of several diseases, especially those involving the cardiovascular system. Sepsis is a complex condition that can lead to multiorgan failure, shock and death. The number of people developing sepsis is still increasing (approximately 750,000 cases of sepsis occur annually in the USA). Both ANP and pro-ANP have attracted interest as new markers for sepsis. Reports indicate that ANP or BNP levels are elevated in septic patients. However, many mechanisms are still unexplained. This situation is complicated by the fact that contradictory results have been published. There are several reasons for this controversy including differences in the techniques used to assay natriuretic peptides. Nevertheless, natriuretic peptides might eventually prove useful for the diagnosis and/or the treatment of septic patients.

Quantitative morphological study of microglial cells in the ischemic rat brain using principal component analysis.

Pathogenic stimuli induce alterations in the morphology of microglial cells. We analysed changes in lectin-stained cells on the 1st, 3rd, 7th or 14th day after transient global ischemia. Three areas differing in the degree of microglial reaction were selected for analysis: the upper cerebral cortex, the hippocampal CA1 area, and the hilus of the dentate gyrus. Nine morphological parameters, including fractal dimension, lacunarity, self-similarity range, solidity, convexity and form factor were determined. Then the resultant data were processed using principal component analysis (PCA). We found that the two first principal components together explained more than 73% of the observed variability, and may be sufficient both to describe the morphological diversity of the cells, and to determine the dynamics and direction of the changes. In both hippocampal areas, the transformation to hypertrophied and phagocytic cells was observed, but changes in the hilus were faster than in the CA1. In contrast, in the cortex, a microglial reaction was characterised by an increase in the complexity of processes. The results presented show that the quantitative morphological analysis can be an effective tool in research on the reactive behaviour of microglia and, particularly, in the detection of small and early changes in the cells.