Bacteremia due to extended-spectrum beta-lactamase-producing Escherichia coli in the CTX-M era: a new clinical challenge.

Extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli, particularly those producing CTX-M types of ESBL, are emerging pathogens. Bacteremia caused by these organisms represents a clinical challenge, because the organisms are frequently resistant to the antimicrobials recommended for treatment of patients with suspected E. coli sepsis. METHODS:A cohort study was performed that included all episodes of bloodstream infection due to ESBL-producing E. coli during the period from January 2001 through March 2005. Data on predisposing factors, clinical presentation, and outcome were collected. ESBLs were characterized using isoelectric focusing, polymerase chain reaction, and sequencing. RESULTS: Forty-three episodes (8.8% of cases of bacteremia due to E. coli) were included; 70% of the isolates produced a CTX-M type of ESBL. The most frequent origins of infection were the urinary (46%) and biliary tracts (21%). Acquisition was nosocomial in 21 cases (49%), health care associated in 14 cases (32%), and strictly community acquired in 8 cases (19%). Thirty-eight percent and 25% of patients had obstructive diseases of the urinary and biliary tracts, respectively, and 38% had recently received antimicrobials. Nine patients (21%) died. Compared with beta-lactam/beta-lactamase-inhibitor and carbapenem-based regimens, empirical therapy with cephalosporins or fluoroquinolones was associated with a higher mortality rate (9% vs. 35%; P=.05) and needed to be changed more frequently (24% vs. 78%; P=.001). CONCLUSIONS: ESBL-producing E. coli is a significant cause of bloodstream infection in hospitalized and nonhospitalized patients in the context of the emergence of CTX-M enzymes. Empirical treatment of sepsis potentially caused by E. coli may need to be reconsidered in areas where such ESBL-producing isolates are present.

Clinical and molecular epidemiology of extended-spectrum beta-lactamase-producing Escherichia coli as a cause of nosocomial infection or colonization: implications for control.

BACKGROUND: Extended-spectrum beta-lactamase (ESBL)-producing members of the Enterobacteriaceae family are important nosocomial pathogens. Escherichia coli producing a specific family of ESBL (the CTX-M enzymes) are emerging worldwide. The epidemiology of these organisms as causes of nosocomial infection is poorly understood. The aims of this study were to investigate the clinical and molecular epidemiology of nosocomial infection or colonization due to ESBL-producing E. coli in hospitalized patients, consider the specific types of ESBLs produced, and identify the risk factors for infection and colonization with these organisms. METHODS: All patients with nosocomial colonization and/or infection due to ESBL-producing E. coli in 2 centers (a tertiary care hospital and a geriatric care center) identified between January 2001 and May 2002 were included. A double case-control study was performed. The clonal relatedness of the isolates was studied by repetitive extragenic palindromic-polymerase chain reaction and pulsed-field gel electrophoresis. ESBLs were characterized by isoelectric focusing, polymerase chain reaction, and sequencing. RESULTS: Forty-seven case patients were included. CTX-M-producing E. coli were clonally unrelated and more frequently susceptible to nonoxyimino-beta-lactams. Alternately, isolates producing SHV- and TEM-type ESBL were epidemic and multidrug resistant. Urinary catheterization was a risk factor for both CTX-M-producing and SHV-TEM-producing isolates. Previous oxyimino-beta-lactam use, diabetes, and ultimately fatal or nonfatal underlying diseases were independent risk factors for infection or colonization with CTX-M-producing isolates, whereas previous fluoroquinolone use was associated with infection or colonization with SHV-TEM-producing isolates. CONCLUSIONS: The epidemiology of ESBL-producing E. coli as a cause of nosocomial infection is complex. Sporadic CTX-M-producing isolates coexisted with epidemic multidrug-resistant SHV-TEM-producing isolates. These data should be taken into account for the design of control measures.

The Presence of Precursors of Benign Pre-B Lymphoblasts (Hematogones) in the Bone Marrow of a Paediatric Patient with Cytomegalovirus Infection

Hematogones are normal B-lymphoid precursors that multiply in the bone marrow of small children and of adults with ferropenic anaemia, neuroblastoma or idiopathic thrombocytopenic purpura. They are not normally found in peripheral blood, and the immunophenotype is virtually indistinguishable from that of B lymphoblasts. We discuss the case of a 3-month infant with an active cytomegalovirus infection, with hepatitis and pancytopenia associated with 13% hematogones in the bone marrow

Decrease in beta-Cell Proliferation Precedes Apoptosis during Diabetes Development in Bio-Breeding/Worcester Rat: Beneficial Role of Exendin-4

In autoimmune type 1 diabetes mellitus, proinflammatory cytokine-mediated apoptosis of beta-cells has been considered to be the first event directly responsible for beta-cell mass reduction. In the Bio-Breeding (BB) rat, an in vivo model used in the study of autoimmune diabetes, beta-cell apoptosis is observed from 9 wk of age and takes place after an insulitis period that begins at an earlier age. Previous studies by our group have shown an antiproliferative effect of proinflammatory cytokines on cultured beta-cells in Wistar rats, an effect that was partially reversed by Exendin-4, an analogue of glucagon-like peptide-1. In the current study, the changes in beta-cell apoptosis and proliferation during insulitis stage were also determined in pancreatic tissue sections in normal and thymectomized BB rats, as well as in Wistar rats of 5, 7, 9, and 11 wk of age. Although stable beta-cell proliferation in Wistar and thymectomized BB rats was observed along the course of the study, a decrease in beta-cell proliferation and beta-cell mass from the age of 5 wk, and prior to the commencement of apoptosis, was noted in BB rats. Exendin-4, in combination with anti-interferon-gamma antibody, induced a near-total recovery of beta-cell proliferation during the initial stages of insulitis. This highlights the importance of early intervention and, as well, the possibilities of new therapeutic approaches in preventing autoimmune diabetes by acting, initially, in the insulitis stage and, subsequently, on beta-cell regeneration and on beta-cell apoptosis.

Incretin receptor null mice reveal key role of GLP-1 but not GIP in pancreatic beta cell adaptation to pregnancy.

Islet adaptations to pregnancy were explored in C57BL6/J mice lacking functional receptors for glucagon-like peptide 1 (GLP-1) and gastric inhibitory polypeptide (GIP). Pregnant wild type mice and GIPRKO mice exhibited marked increases in islet and beta cell area, numbers of medium/large sized islets, with positive effects on Ki67/Tunel ratio favouring beta cell growth and enhanced pancreatic insulin content. Alpha cell area and glucagon content were unchanged but prohormone convertases PC2 and PC1/3 together with significant amounts of GLP-1 and GIP were detected in alpha cells. Knockout of GLP-1R abolished these islet adaptations and paradoxically decreased pancreatic insulin, GLP-1 and GIP. This was associated with abolition of normal pregnancy-induced increases in plasma GIP, L-cell numbers, and intestinal GIP and GLP-1 stores. These data indicate that GLP-1 but not GIP is a key mediator of beta cell mass expansion and related adaptations in pregnancy, triggered in part by generation of intra-islet GLP-1.

Phylogenetic alpha and beta diversities of butterfly communities correlate with climate in the western Swiss Alps

The observation of non-random phylogenetic distribution of traits in communities provides evidence for niche-based community assembly. Environment may influence the phylogenetic structure of communities because traits determining how species respond to prevailing conditions can be phylogenetically conserved. In this study, we investigate the variation of butterfly species richness and of phylogenetic - and -diversities along temperature and plant species richness gradients. Our study indicates that butterfly richness is independently positively correlated to temperature and plant species richness in the study area. However, the variation of phylogenetic - and -diversities is only correlated to temperature. The significant phylogenetic clustering at high elevation suggests that cold temperature filters butterfly lineages, leading to communities mostly composed of closely related species adapted to those climatic conditions. These results suggest that in colder and more severe conditions at high elevations deterministic processes and not purely stochastic events drive the assemblage of butterfly communities.

Timing of adequate antibiotic therapy is a greater determinant of outcome than are TNF and IL-10 polymorphisms in patients with sepsis.

INTRODUCTION Genetic variations may influence clinical outcomes in patients with sepsis. The present study was conducted to evaluate the impact on mortality of three polymorphisms after adjusting for confounding variables, and to assess the factors involved in progression of the inflammatory response in septic patients. METHOD The inception cohort study included all Caucasian adults admitted to the hospital with sepsis. Sepsis severity, microbiological information and clinical variables were recorded. Three polymorphisms were identified in all patients by PCR: the tumour necrosis factor (TNF)-alpha 308 promoter polymorphism; the polymorphism in the first intron of the TNF-beta gene; and the IL-10-1082 promoter polymorphism. Patients included in the study were followed up for 90 days after hospital admission. RESULTS A group of 224 patients was enrolled in the present study. We did not find a significant association among any of the three polymorphisms and mortality or worsening inflammatory response. By multivariate logistic regression analysis, only two factors were independently associated with mortality, namely Acute Physiology and Chronic Health Evaluation (APACHE) II score and delayed initiation of adequate antibiotic therapy. In septic shock patients (n = 114), the delay in initiation of adequate antibiotic therapy was the only independent predictor of mortality. Risk factors for impairment in inflammatory response were APACHE II score, positive blood culture and delayed initiation of adequate antibiotic therapy. CONCLUSION This study emphasizes that prompt and adequate antibiotic therapy is the cornerstone of therapy in sepsis. The three polymorphisms evaluated in the present study appear not to influence the outcome of patients admitted to the hospital with sepsis.

Altered expression of beta-galactosidase-1-like protein 3 (Glb1l3) in the retinal pigment epithelium (RPE)-specific 65-kDa protein knockout mouse model of Leber's congenital amaurosis

Purpose: In this study, we investigated the expression of the gene encoding beta-galactosidase (Glb)-1-like protein 3 (Glb1l3), a member of the glycosyl hydrolase 35 family, during retinal degeneration in the retinal pigment epithelium (RPE)-specific 65-kDa protein knockout (Rpe65(-/-)) mouse model of Leber congenital amaurosis (LCA). Additionally, we assessed the expression of the other members of this protein family, including beta-galactosidase-1 (Glb1), beta-galactosidase-1-like (Glb1l), and beta-galactosidase-1-like protein 2 (Glb1l2).Methods: The structural features of Glb1l3 were assessed using bioinformatic tools. mRNA expression of Glb-related genes was investigated by oligonucleotide microarray, real-time PCR, and reverse transcription (RT) -PCR. The localized expression of Glb1l3 was assessed by combined in situ hybridization and immunohistochemistry.Results: Glb1l3 was the only Glb-related member strongly downregulated in Rpe65(-/-) retinas before the onset and during progression of the disease. Glb1l3 mRNA was only expressed in the retinal layers and the RPE/choroid. The other Glb-related genes were ubiquitously expressed in different ocular tissues, including the cornea and lens. In the healthy retina, expression of Glb1l3 was strongly induced during postnatal retinal development; age-related increased expression persisted during adulthood and aging.Conclusions: These data highlight early-onset downregulation of Glb1l3 in Rpe65-related disease. They further indicate that impaired expression of Glb1l3 is mostly due to the absence of the chromophore 11-cis retinal, suggesting that Rpe65 deficiency may have many metabolic consequences in the underlying neuroretina.

GLP-1 protects beta-cells against apoptosis by enhancing the activity of an IGF-2/IGF-1 receptor autocrine loop

GLP-1 protects β-cells against apoptosis by still incompletely understood mechanisms. In a recent study, we searched for novel anti-apoptotic pathways by performing comparative transcriptomic analysis of islets from Gipr-/-;Glp-1r-/- mice, which show increased susceptibility to cytokine-induced apoptosis. We observed a strong reduction in IGF-1R expression in the knockout islets suggesting a link between the gluco-incretin and IGF-1R signaling pathways. Using MIN6 and primary islet cells, we demonstrated that GLP-1 strongly stimulates IGF-1R expression and that activation of the IGF-1R/Akt signaling pathway required active secretion of IGF-2 by the β-cells. We showed that inactivation of the IGF-1 receptor gene in β-cells or preventing its up-regulation by GLP-1, as well as suppressing IGF-2 expression or action, blocked the protective effect of GLP-1 against cytokine-induced apoptosis. Thus, an IGF-2/IGF-1 receptor autocrine loop operates in β-cells and GLP-1 increases its activity by enhancing IGF-1R expression and by stimulating IGF-2 secretion. This mechanism is required for GLP-1 to protect β-cells against apoptosis.

Efficient generation of human alloantigen-specific CD4+ regulatory T cells from naive precursors by CD40-activated B cells.

CD4(+)CD25(+)Foxp3(+) regulatory T cells (Treg) play an important role in the induction and maintenance of immune tolerance. Although adoptive transfer of bulk populations of Treg can prevent or treat T cell-mediated inflammatory diseases and transplant allograft rejection in animal models, optimal Treg immunotherapy in humans would ideally use antigen-specific rather than polyclonal Treg for greater specificity of regulation and avoidance of general suppression. However, no robust approaches have been reported for the generation of human antigen-specific Treg at a practical scale for clinical use. Here, we report a simple and cost-effective novel method to rapidly induce and expand large numbers of functional human alloantigen-specific Treg from antigenically naive precursors in vitro using allogeneic nontransformed B cells as stimulators. By this approach naive CD4(+)CD25(-) T cells could be expanded 8-fold into alloantigen-specific Treg after 3 weeks of culture without any exogenous cytokines. The induced alloantigen-specific Treg were CD45RO(+)CCR7(-) memory cells, and had a CD4(high), CD25(+), Foxp3(+), and CD62L (L-selectin)(+) phenotype. Although these CD4(high)CD25(+)Foxp3(+) alloantigen-specific Treg had no cytotoxic capacity, their suppressive function was cell-cell contact dependent and partially relied on cytotoxic T lymphocyte antigen-4 expression. This approach may accelerate the clinical application of Treg-based immunotherapy in transplantation and autoimmune diseases.

HLA Alleles Influence the Clinical Signature of Amoxicillin-Clavulanate Hepatotoxicity.

BACKGROUND AND AIM The genotype-phenotype interaction in drug-induced liver injury (DILI) is a subject of growing interest. Previous studies have linked amoxicillin-clavulanate (AC) hepatotoxicity susceptibility to specific HLA alleles. In this study we aimed to examine potential associations between HLA class I and II alleles and AC DILI with regards to phenotypic characteristics, severity and time to onset in Spanish AC hepatotoxicity cases. METHODS High resolution genotyping of HLA loci A, B, C, DRB1 and DQB1 was performed in 75 AC DILI cases and 885 controls. RESULTS The distributions of class I alleles A*3002 (P/Pc = 2.6E-6/5E-5, OR 6.7) and B*1801 (P/Pc = 0.008/0.22, OR 2.9) were more frequently found in hepatocellular injury cases compared to controls. In addition, the presence of the class II allele combination DRB1*1501-DQB1*0602 (P/Pc = 5.1E-4/0.014, OR 3.0) was significantly increased in cholestatic/mixed cases. The A*3002 and/or B*1801 carriers were found to be younger (54 vs 65 years, P = 0.019) and were more frequently hospitalized than the DRB1*1501-DQB1*0602 carriers. No additional alleles outside those associated with liver injury patterns were found to affect potential severity as measured by Hy's Law criteria. The phenotype frequencies of B*1801 (P/Pc = 0.015/0.42, OR 5.2) and DRB1*0301-DQB1*0201 (P/Pc = 0.0026/0.07, OR 15) were increased in AC DILI cases with delayed onset compared to those corresponding to patients without delayed onset, while the opposite applied to DRB1*1302-DQB1*0604 (P/Pc = 0.005/0.13, OR 0.07). CONCLUSIONS HLA class I and II alleles influence the AC DILI signature with regards to phenotypic expression, latency presentation and severity in Spanish patients.

A pilot study of delayed versus immediate serial casting after botulinum toxin injection for partially reducible spastic equinus.

BACKGROUND: Serial casting is often prescribed after botulinum toxin injections to improve joint ranges of motion and to potentiate the decrease in hypertonia. The aim of this study was to compare delayed versus immediate serial casting as an adjunct to botulinum toxin therapy for partially reducible spastic equinus. METHODS: Twelve children who presented spastic equinus associated with mild gastrosoleus contracture took part. Five of them had a diagnosis of spastic diplegia, whereas 7 had a diagnosis of congenital hemiplegia. Children were randomized to immediate serial casting (same day) or delayed serial casting (4 weeks later) after botulinum toxin injection to their gastrosolei. Casts were replaced weekly for 3 weeks. RESULTS: Three children complained of pain that required recasting in the immediate casting group versus none in the delayed casting group (P = 0.08). At 3 months, there was a 27-degree improvement in the fast dorsiflexion angle (Tardieu R1) in the delayed casting group versus 17 degrees in the immediate casting group (P = 0.029). At 6 months, a 19-degree improvement persisted in the delayed group compared with 11 degrees in the immediate group (P = 0.010). CONCLUSIONS: There is a clear benefit in delaying serial casting after the injection of botulinum toxin in the recurrence of spasticity at the gastrosoleus that may also offer an advantage regarding the incidence of painful episodes associated with casting. Most importantly, reducing the recurrence of spasticity by delayed serial casting may offer the possibility of decreasing the frequency of botulinum toxin reinjections.

The scaffold protein IB1/JIP-1 is a critical mediator of cytokine-induced apoptosis in pancreatic beta cells.

In insulin-secreting cells, cytokines activate the c-Jun N-terminal kinase (JNK), which contributes to a cell signaling towards apoptosis. The JNK activation requires the presence of the murine scaffold protein JNK-interacting protein 1 (JIP-1) or human Islet-brain 1(IB1), which organizes MLK3, MKK7 and JNK for proper signaling specificity. Here, we used adenovirus-mediated gene transfer to modulate IB1/JIP-1 cellular content in order to investigate the contribution of IB1/JIP-1 to beta-cell survival. Exposure of the insulin-producing cell line INS-1 or isolated rat pancreatic islets to cytokines (interferon-gamma, tumor necrosis factor-alpha and interleukin-1beta) induced a marked reduction of IB1/JIP-1 content and a concomitant increase in JNK activity and apoptosis rate. This JNK-induced pro-apoptotic program was prevented in INS-1 cells by overproducing IB1/JIP-1 and this effect was associated with inhibition of caspase-3 cleavage. Conversely, reducing IB1/JIP-1 content in INS-1 cells and isolated pancreatic islets induced a robust increase in basal and cytokine-stimulated apoptosis. In heterozygous mice carrying a selective disruption of the IB1/JIP-1 gene, the reduction in IB1/JIP-1 content in happloinsufficient isolated pancreatic islets was associated with an increased JNK activity and basal apoptosis. These data demonstrate that modulation of the IB1-JIP-1 content in beta cells is a crucial regulator of JNK signaling pathway and of cytokine-induced apoptosis.

Candidate gene study of TRAIL and TRAIL receptors: association with response to interferon beta therapy in multiple sclerosis patients.

TRAIL and TRAIL Receptor genes have been implicated in Multiple Sclerosis pathology as well as in the response to IFN beta therapy. The objective of our study was to evaluate the association of these genes in relation to the age at disease onset (AAO) and to the clinical response upon IFN beta treatment in Spanish MS patients. We carried out a candidate gene study of TRAIL, TRAILR-1, TRAILR-2, TRAILR-3 and TRAILR-4 genes. A total of 54 SNPs were analysed in 509 MS patients under IFN beta treatment, and an additional cohort of 226 MS patients was used to validate the results. Associations of rs1047275 in TRAILR-2 and rs7011559 in TRAILR-4 genes with AAO under an additive model did not withstand Bonferroni correction. In contrast, patients with the TRAILR-1 rs20576-CC genotype showed a better clinical response to IFN beta therapy compared with patients carrying the A-allele (recessive model: p = 8.88×10(-4), pc = 0.048, OR = 0.30). This SNP resulted in a non synonymous substitution of Glutamic acid to Alanine in position 228 (E228A), a change previously associated with susceptibility to different cancer types and risk of metastases, suggesting a lack of functionality of TRAILR-1. In order to unravel how this amino acid change in TRAILR-1 would affect to death signal, we performed a molecular modelling with both alleles. Neither TRAIL binding sites in the receptor nor the expression levels of TRAILR-1 in peripheral blood mononuclear cell subsets (monocytes, CD4+ and CD8+ T cells) were modified, suggesting that this SNP may be altering the death signal by some other mechanism. These findings show a role for TRAILR-1 gene variations in the clinical outcome of IFN beta therapy that might have relevance as a biomarker to predict the response to IFN beta in MS.

Trans-scleral local resection of toxic choroidal melanoma after proton beam radiotherapy.

AIM: To report on trans-scleral local resection of choroidal melanoma for exudative retinal detachment and neovascular glaucoma (toxic tumour syndrome) after proton beam radiotherapy (PBR). METHODS: A non-randomised, prospective study of secondary trans-scleral local resection of choroidal melanoma for exudative retinal detachment with or without neovascular glaucoma after PBR. The patients were treated at the Liverpool Ocular Oncology Centre between February 2000 and April 2008. The trans-scleral local resection was performed with a lamellar-scleral flap, using systemic hypotension to reduce haemorrhage. RESULTS: 12 patients (six women, six men) with a mean age of 51 years (range 20-75) were included in this study. The tumour margins extended anterior to ora serrata in six patients. On ultrasonography, the largest basal tumour dimension averaged 12.4 mm (range 6.8-18.1) and the tumour height averaged 7.1 mm (range 4.2-10.7). The retinal detachment was total in seven patients. Neovascular glaucoma was present in four patients. The time between PBR and local resection had a mean of 17.4 months (range 1-84). The ophthalmic follow-up time after the local resection had a mean of 46.2 months (range 14-99). At the latest known status, the eye was conserved in 10 patients, with a flat retina in all these patients and visual acuity equal or better than 6/30 in four patients. The reasons for enucleation were: patient request for enucleation when rhegmatogenous retinal detachment complicated the resection (one patient) and phthisis (one patient). CONCLUSIONS: Exudative retinal detachment, rubeosis and neovascular glaucoma after PBR of a choroidal melanoma can resolve after trans-scleral local resection of the tumour. Our findings suggest that these complications are caused by the persistence of the irradiated tumour within the eye ('toxic tumour syndrome').