Bordetella bronchiseptica are small, pleomorphic Gram-negative coccobacilli which are commensal organisms in the upper respiratory tract of many wild and domestic animals ('kennel cough' in dogs). While it is common for health care providers to ask about exposure to ill family/friends, most do not routinely inquire about the health or immunization status of household pets. We report two cases of B. bronchiseptica pneumonia in lung transplant recipients [cystic fibrosis (CF); ages 10 and 15 yr; one male] who contracted B. bronchiseptica from pet dogs. We compared their course and outcome to four children (two CF, one congenital heart disease and one Duchenne's muscular dystrophy; four males, age range 6 months to 14 yr) with B. bronchiseptica cultured from the respiratory tract. Two of the four patients also acquired their illnesses from pet dogs and two from unknown sources. One lung transplant recipient expired from progressive respiratory failure. We conclude that B. bronchiseptica can cause serious infections in both immunosuppressed and immunocompetent children. We speculate that a detailed history of exposure to ill pets (particularly dogs), and the immunization status of all pets should be included in the routine evaluation of all pediatric transplant recipients.

A normative framework for modeling causal and counterfactual reasoning has been proposed by Spirtes, Glymour, and Scheines (1993; cf. Pearl, 2000). The framework takes as fundamental that reasoning from observation and intervention differ. Intervention includes actual manipulation as well as counterfactual manipulation of a model via thought. To represent intervention, Pearl employed the do operator that simplifies the structure of a causal model by disconnecting an intervened-on variable from its normal causes. Construing the do operator as a psychological function affords predictions about how people reason when asked counterfactual questions about causal relations that we refer to as undoing, a family of effects that derive from the claim that intervened-on variables become independent of their normal causes. Six studies support the prediction for causal (A causes B) arguments but not consistently for parallel conditional (if A then B) ones. Two of the studies show that effects are treated as diagnostic when their values are observed but nondiagnostic when they are intervened on. These results cannot be explained by theories that do not distinguish interventions from other sorts of events.

OBJECTIVE

The loss of pancreatic beta-cells is thought to be one of the principal causes of diabetes mellitus (DM) in cystic fibrosis (CF), but the role of peripheral insulin resistance (IR) in the pathogenesis of DM in CF remains unclear. The aim of this study was to evaluate whether eventual changes of glucose tolerance (GT) over time were associated with modifications of insulin secretion or sensitivity.

METHODS

Plasma glucose and insulin responses to an oral GT test (OGTT) were investigated and reinvestigated 13 Years later in 14 CF patients with initial and persistent fasting euglycemia and no history of insulin treatment. Insulin sensitivity (IS) at both tests was assessed on the basis of insulin and glucose levels both in the fasting state and during OGTTs.

RESULTS

From the 1st to the 2nd OGTT: (a) the prevalence of DM responses significantly increased; (b) the areas beneath the respective glucose and insulin curves significantly increased and decreased respectively; (c) IR and IS indices decreased and increased respectively, even in the patients who developed DM; (d) pulmonary function significantly worsened in the entire series, especially in the patients who developed DM.

CONCLUSIONS

(i) the natural history of glyco-metabolic status in CF is characterized by deteriorating GT over time; (ii) insulinopenia plays a prominent role in the pathogenesis of GT worsening; (iii) IR does not play any significant part in the pathogenesis of DM development; (iv) deterioration of lung function tests is more severe in the subjects who develop DM over time.

Burkholderia cenocepacia is an opportunistic pathogen that primarily infects cystic fibrosis (CF) patients. Previously, we reported that ShvR, a LysR regulator, influences colony morphology, virulence, and biofilm formation and regulates the expression of an adjacent 24-kb genomic region encoding 24 genes. In this study, we report the functional characterization of selected genes in this region. A Tn5 mutant with shiny colony morphology was identified with a polar mutation in BCAS0208, predicted to encode an acyl-coenzyme A dehydrogenase. Mutagenesis of BCAS0208 and complementation analyses revealed that BCAS0208 is required for rough colony morphology, biofilm formation, and virulence on alfalfa seedlings. It was not possible to complement with BCAS0208 containing a mutation in the catalytic site. BCAS0201, encoding a putative flavin adenine dinucleotide (FAD)-dependent oxidoreductase, and BCAS0207, encoding a putative citrate synthase, do not influence colony morphology but are required for optimum levels of biofilm formation and virulence. Both BCAS0208 and BCAS0201 contribute to pellicle formation, although individual mutations in each of these genes had no appreciable effect on pellicle formation. A mutant with a polar insertion in BCAS0208 was significantly less virulent in a rat model of chronic lung infection as well as in the alfalfa model. Genes in this region were shown to influence utilization of branched-chain fatty acids, tricarboxylic acid cycle substrates, l-arabinose, and branched-chain amino acids. Together, our data show that the ShvR-regulated genes BCAS0208 to BCAS0201 are required for the rough colony morphotype, biofilm and pellicle formation, and virulence in B. cenocepacia.

The release of "neutrophil extracellular traps" (NETs) has been identified as a novel immune response in innate immunity. NETs are composed of neutrophil-derived circulating free DNA (cf-DNA) and neutrophil cytoplasm-derived proteins such as proteases. In this study, we analyzed the putative diagnostic value of synovial cf-DNA/NETs for identification of septic arthritis. Forty-two patients with a joint effusion who had undergone arthrocentesis were included. From synovial fluid, cf-DNA/NETs (j-cf-DNA) levels were directly quantified. Diagnostic value of j-cf-DNA was compared with white blood cells (WBC), synovial white blood cells (j-WBC), C-reactive protein (CRP), j-IL-6, j-TNF alpha, j-IL-1 beta, and myeloperoxidase (j-MPO). Sensitivity, specificity, positive and negative predictive value, as well as ROC-curves for each parameter were calculated. Synovial fluid cf-DNA/NETs values from patients with septic arthritis (3,286 +/- 386 ng/ml, n = 9) were significantly increased compared to patients with noninfectious joint inflammation (1,040 +/- 208 ng/ml, n = 17) or osteoarthritis (278 +/- 34 ng/ml, n = 16, p < 0.01). In conjunction with j-cf-DNA, j-IL-6 and j-IL-1 beta were significantly elevated (p < 0.01), but WBC, CRP, and j-WBC were not. At a cut-off of 300 ng/ml, j-cf-DNA had a sensitivity of 0.89, a specificity of 1.0, a positive predictive value of 1.0, and a negative predictive value of 0.97. Receiver operation curves revealed largest areas under the curve for cf-DNA/NETs (0.933) and j-IL-6 (0.951). cf-DNA/NETs seem to be a valuable additional marker for the diagnosis of septic arthritis or periprosthetic infections. However, this result should be confirmed in a large clinical trial.

B. cenocepacia is an opportunistic human pathogen that is particularly problematic for patients suffering from cystic fibrosis (CF). In the CF lung bacteria grow to high densities within the viscous mucus that is limited in oxygen. Pseudomonas aeruginosa, the dominant pathogen in CF patients, is known to grow and survive under oxygen-limited to anaerobic conditions by using micro-oxic respiration, denitrification and fermentative pathways. In contrast, inspection of the genome sequences of available B. cenocepacia strains suggested that B. cenocepacia is an obligate aerobic and non-fermenting bacterium. In accordance with the bioinformatics analysis we observed that B. cenocepacia H111 is able to grow with as little as 0.1% O2 but not under strictly anoxic conditions. Phenotypic analyses revealed that H111 produced larger amounts of biofilm, pellicle and proteases under micro-oxic conditions (0.5%-5% O2, i.e. conditions that mimic those encountered in CF lung infection), and was more resistant to several antibiotics. RNA-Seq and shotgun proteomics analyses of cultures of B. cenocepacia H111 grown under micro-oxic and aerobic conditions showed up-regulation of genes involved in the synthesis of the exopolysaccharide (EPS) cepacian as well as several proteases, two isocitrate lyases and other genes potentially important for life in micro-oxia.

UNASSIGNED

RNA-Seq raw data files are accessible through the GEO Series accession number GSE48585. MS data have been deposited in the ProteomeXchange database (PXD000270).

Pharmacodynamic data on ciprofloxacin indicate that a target area under the concentration-time curve from 0 to 24 h (AUC(0-24))/MIC ratio of>>or=125 is necessary to achieve optimal bactericidal activity for the treatment of gram-negative pneumonia. The purpose of this prospective study was to (i) develop a pharmacokinetic (PK) model to be utilized for therapeutic drug monitoring (TDM) of ciprofloxacin and (ii) evaluate current ciprofloxacin dosing regimens for pneumonias in cystic fibrosis (CF) patients. Twelve adult CF patients received a single 400-mg dose of IV ciprofloxacin. Six blood samples were obtained over a 12-h interval. Serum drug concentrations were determined by high-pressure liquid chromotography and were fitted to one- and two-compartment models by using NPEM2. Ciprofloxacin MIC data for Pseudomonas aeruginosa were obtained from 1,213 CF patients enrolled in a large clinical trial. A Monte Carlo simulation was performed to estimate the fractional attainment of an AUC(0-24)/MIC ratio of>>or=125. A two-compartment model best describes the serum drug concentration data. The mean fitted PK parameter values are volume of distribution in the central compartment, 0.29 liter/kg; volume of distribution at steady state, 1.1 liters/kg; total clearance, 0.34 liter/h/kg; distributional clearance, 0.89 liter/h/kg; half-life at alpha phase, 0.16 h; and half-life at beta phase, 2.9 h. The overall fractional attainment of achieving an AUC(0-24)/MIC ratio of>>or=125 against P. aeruginosa isolates with ciprofloxacin (400 mg every 12 h [q12h] and 8 qh) were 10 and 30%, respectively. A clinical breakpoint MIC of <0.5 microg/ml for susceptibility is suggested, based on an examination of the fractional attainment of the AUC(0-24)/MIC target at each MIC. The recommended doses of 400 mg q8h or q12h may be inadequate to treat an acute pulmonary exacerbation when given alone. The poor and variable AUC(0-24)/MIC ratios support the use of TDM to monitor and adjust the dosage to optimize the efficacy of ciprofloxacin therapy in these patients.

Diffusion coefficients for the intercellular movement of fluorescent tracers have been measured in the epidermis of a larval beetle. Fluorescent tracer was injected into a cell and the spread of tracer from cell to cell in this monolayer was recorded by a TV camera. Fluorescence intensities were digitized from the TV images at successive times after the start of injection at various distances from the source by a microcomputer interfaced with a video analyzer. From the relationship between concentration (measured as light intensity), time and distance, an effective diffusion coefficient (De) is calculated for the tracer in the tissue. In newly ecdysed epidermis, De for carboxyfluorescein (CF) is 2.7 X 10(-7) cm2/s, and De for lissamine rhodamine B (LRB) is 1.2 X 10(-7) cm2/s, whereas in intermolt epidermis the De's for CF and LRB are 3.7 X 10(-7) and 1.2 X 10(-7) cm2/s, respectively. These diffusion coefficients are only an order of magnitude lower than their values in water. The ratio of De for the two tracers at these two stages of development differs from the ratio predicted in cytoplasm alone, with the movement of the slightly larger molecule (LRB) being impeded relative to that of the smaller molecule (CF). This suggests that the properties of the membrane channels amplify differences in the rates of movement of molecules of similar size. This may be important during cell patterning in development. De for CF was also monitored as junctional resistance was increased in the epidermis. During 30 min of exposure to 0.25 mM chlorpromazine, De dropped to 20% of its initial value of 5 X 10(-7) cm2/s, implying that the junctional membrane, rather than cytoplasm, is the major barrier to molecular diffusion among the cells.

BACKGROUND

A multi-centre, four-arm trial (the PACE trial) found that rehabilitative cognitive behaviour therapy (CBT) and graded exercise therapy (GET) were more effective treatments for chronic fatigue syndrome (CFS) than specialist medical care (SMC) alone, when each was added to SMC, and more effective than adaptive pacing therapy (APT) when added to SMC. In this study we compared how many participants recovered after each treatment.

METHODS

We defined recovery operationally using multiple criteria, and compared the proportions of participants meeting each individual criterion along with two composite criteria, defined as (a) recovery in the context of the trial and (b) clinical recovery from the current episode of the illness, however defined, 52 weeks after randomization. We used logistic regression modelling to compare treatments.

RESULTS

The percentages (number/total) meeting trial criteria for recovery were 22% (32/143) after CBT, 22% (32/143) after GET, 8% (12/149) after APT and 7% (11/150) after SMC. Similar proportions met criteria for clinical recovery. The odds ratio (OR) for trial recovery after CBT was 3.36 [95% confidence interval (CI) 1.64–6.88] and for GET 3.38 (95% CI 1.65–6.93), when compared to APT, and after CBT 3.69 (95% CI 1.77–7.69) and GET 3.71 (95% CI 1.78–7.74), when compared to SMC (p values < or =0.001 for all comparisons). There was no significant difference between APT and SMC. Similar proportions recovered in trial subgroups meeting different definitions of the illness.

CONCLUSIONS

This study confirms that recovery from CFS is possible, and that CBT and GET are the therapies most likely to lead to recovery.

The clinical course of cystic fibrosis (CF) varies considerably among patients carrying the same CF-causing gene mutation. Additional genetic modifiers may contribute to this variability. As airway inflammation is a key component of CF pathophysiology, we investigated whether major cytokine variants represent such modifiers in young CF patients. We tested 13 polymorphisms in 8 genes that play a key role in the inflammatory response: tumor necrosis factor, lymphotoxin alpha, interleukin (IL) 1B, IL1 receptor antagonist, IL6, IL8, IL10 and transforming growth factor beta 1 (TGFB1), for an association with lung disease progression and nutritional status in 329 CF patients. Variants in the TGFB1 gene at position +869T/C demonstrated a significant association with lung function decline. A less pronounced rate of decline in forced expiratory volume in 1 sec (FEV(1)) and forced vital capacity (FVC) were observed in patients heterozygous for TGFB1 +869 (+869CT), when compared to patients carrying either TGFB1 +869TT or +869CC genotypes. These findings support the concept that TGFB1 gene variants appear to be important genetic modifiers of lung disease progression in CF.

Immune adherence hemagglutination (IAHA) was compared to complement fixation (CF), using standard procedures, for serological testing of human sera with a number of commercially available antigens. The antigens included herpes simplex, measles, cytomegalo-, and influenza (type B) viruses, as well as Mycoplasma pneumoniae and Chlamydia psittaci (Chlamydia group). The IAHA test was found to be as specific as the CF test, but 4 to 20 times as sensitive with all antigens tested. Antigen titers were also higher with the IAHA method, and the time required to complete the test was only 4 h for the IAHA method, compared with 20 h for the CF method. The increased sensitivity of the IAHA test should permit its use for determination of immunity, as well as for serodiagnosis of recent infections.

Cerebrospinal fluid (CSF) and serum from 35 pairs of multiple sclerosis (MS) patients were analysed as regards mononuclear pleocytosis, concentrations of total protein, immunoglobulin G and A and beta-trace protein, and kappa:lambda ratios, as well as the serum/CSF ratios of IgG and albumin. The disability of the patients differed, whereas the age and the duration of the disease were similar in each pair. Similar analyses were also performed on CSF and serum from 72 patients, who were subdivided according to age at onset and severity of the disease. The highest mean values of the CSF-IgG and the lowest mean values of the serum/CSF IgG ratios were found in the more disabled patients. CSF immunoglobulin abnormalities were encountered more often and were more pronounced in the patients with the most malignant course of the disease, i.e., in those with severe disability after a short duration of the disease (less than 10 yr) and in severely disabled patients with an early age at onset of the disease(less than 25 yr). Contrarily, normal mean values of CSF-IgG concentrations and serum/CFS/IgG ratios were found in the groups of patients without disability after a duration of the disease of 10 years or more, and patients without disability and an early age at onset of the disease (less than 25 yr). The observations indicate that the immune response is most vigorous in disabled patients with a short duration or with an early age at onset of the disease. MS patients with a late age at onset (greater than 35 yr) showed a less pronouced immune response within the CNS, irrespective of the occurrence of disability. The most disabled patients also showed the most severe blood-brain barrier damage as manifested by high mean values of total protein in CSF and low serum/CSF albumin ratios. The patients with severe disability and a long duration of the disease (greater than 10 yr) had the highest content of beta-trace protein in the CSF, probably as a sign of destruction of brain matter.

The molecular pathogenesis of cystic fibrosis (CF) liver disease is unknown. This study investigates its earliest pathophysiological manifestations employing a mouse model carrying DeltaF508, the commonest human CF mutation. We hypothesized that, if increased bile salt spillage into the colon occurs as in the human disease, then this should lead to a hydrophobic bile salt profile and to "hyperbilirubinbilia" because of induced enterohepatic cycling of unconjugated bilirubin. Hyperbilirubinbilia may then lead to an increased bile salt-to-phospholipid ratio in bile and, following hydrolysis, precipitation of divalent metal salts of unconjugated bilirubin. We document in CF mice elevated fecal bile acid excretion and biliary secretion of more hydrophobic bile salts compared with control wild-type mice. Biliary secretion rates of bilirubin monoglucuronosides, bile salts, phospholipids, and cholesterol are increased significantly with an augmented bile salt-to-phospholipid ratio. Quantitative histopathology of CF livers displays mild early cholangiopathy in approximately 53% of mice and multifocal divalent metal salt deposition in cholangiocytes. We conclude that increased fecal bile acid loss leads to more hydrophobic bile salts in hepatic bile and to hyperbilirubinbilia, a major contributor in augmenting the bile salt-to-phospholipid ratio and endogenous beta-glucuronidase hydrolysis of bilirubin glucuronosides. The confluence of these perturbations damages intrahepatic bile ducts and facilitates entrance of unconjugated bilirubin into cholangiocytes. This study of the earliest stages of CF liver disease provides a framework for investigating the molecular pathophysiology of more advanced disease in murine models and in humans with CF.

It was proposed that the cystic fibrosis transmembrane conductance regulator (CFTR) functions in the endosomal compartment as a adenosine 3',5'-cyclic monophosphate (cAMP)-regulated Cl channel that regulates endosomal acidification (J. Barasch, B. Kiss, A. Prince, L. Saiman, D. Gruenert, and A. Al-Awqati, Nature Lond. 352: 70-73, 1991). This hypothesis was tested in stably transfected Swiss 3T3 fibroblasts expressing CFTR or delta F508 CFTR and in T84 epithelial cells that normally express CFTR. In fibroblasts, the time course of pH in individual endosomes was measured by quantitative image analysis after 1 min pulse labeling with 2 microM carboxyfluorescein (Cf)-tetramethylrhodamine-transferrin (K. Zen, J. Biwersi, N. Periasamy, and A. S. Verkman. J. Cell Biol. 119: 99-110, 1992). Average endosomal pH reached 6.20 +/- 0.07 (SE) after 15 min in the mock-transfected cells with a half time of approximately 3 min; pH was slightly lower (5.97 +/- 0.06) in the CFTR-expressing fibroblasts. The difference did not result from a subpopulation of highly acidic endosomes. Forskolin (10 microM) increased average pH to 6.62 +/- 0.03 and abolished the difference. For determination of Cl conductance, endosomes in fibroblasts and T84 cells were labeled with Cf-dextran (5 mg/ml); dissipation of the endosomal pH gradient was measured in response to rapid addition of the protonophore carbonyl cyanide m-chlorophenylhydrazone (CCCP; 20 microM). Because the proton flux across the endosomal membrane is limited by the movement of K and Cl, the rate of alkalinization (dpH/dt) after CCCP addition provided a measure of endosomal Cl conductance. In CFTR-expressing fibroblasts, forskolin (10 microM) increased dpH/dt 1.6 +/- 0.2-fold (n = 14).(ABSTRACT TRUNCATED AT 250 WORDS)

Chronic fatigue syndrome (CFS) is a heterogeneous disorder of unknown aetiology characterized by disabling fatigue, headaches, sleep disturbance and several other symptoms. The onset of CFS may follow a viral infection or period of stress. Patients with CFS do not have hypogammaglobulinaemia, predisposition to recurrent bacterial infections or symptoms of autoimmunity. To date, defects in B cell numbers or function have not been shown in the literature. However, treatment with anti-B cell therapy using Rituximab has recently shown benefit to CFS patients. We therefore postulated that patients with CFS had a subtle humoral immune dysfunction, and performed extended B cell immunophenotyping. We undertook a detailed characterization of the proportions of the different B cell subsets in 33 patients with CFS fulfilling the Canadian and Fukada criteria for CFS and compared these with 24 age- and gender-matched healthy controls (HC). CFS patients had greater numbers of naive B cells as a percentage of lymphocytes: 6·3 versus 3·9% in HC (P = 0·034), greater numbers of naive B cells as a percentage of B cells: 65 versus 47% in controls (P = 0·003), greater numbers of transitional B cells: 1·8 versus 0·8% in controls (P = 0·025) and reduced numbers of plasmablasts: 0·5 versus 0·9% in controls (P = 0·013). While the cause of these changes is unclear, we speculate whether they may suggest a subtle tendency to autoimmunity.

Despite considerable worldwide efforts, no single etiology has been identified to explain the development of chronic fatigue syndrome (CFS). It is likely that multiple factors promote its development, sometimes with the same factors both causing and being caused by the syndrome. A detailed review of the literature suggests a number of marginal nutritional deficiencies may have etiologic relevance. These include deficiencies of various B vitamins, vitamin C, magnesium, sodium, zinc, L-tryptophan, L-carnitine, coenzyme Q10, and essential fatty acids. Any of these nutrients could be marginally deficient in CFS patients, a finding that appears to be primarily due to the illness process rather than to inadequate diets. It is likely that marginal deficiencies not only contribute to the clinical manifestations of the syndrome, but also are detrimental to the healing processes. Therefore, when feasible, objective testing should identify them and their resolution should be assured by repeat testing following initiation of treatment. Moreover, because of the rarity of serious adverse reactions, the difficulty in ruling out marginal deficiencies, and because some of the therapeutic benefits of nutritional supplements appear to be due to pharmacologic effects, it seems rational to consider supplementing CFS patients with the nutrients discussed above, along with a general high-potency vitamin/mineral supplement, at least for a trial period.

The ability of ethyl alcohol to modify responses to stress has been well documented (cf. Pohorecky, 1990). However, the structural substrate mediating these effects of alcohol remains undefined. Using immediate early gene (IEG) expression in the brain as a marker of altered neuronal response, we investigated the effect of acute alcohol exposure on the activity of brain regions of rats exposed to 15 min of restraint stress. Immunocytochemical localization c-Fos protein demonstrated that restraint stress led to an induction of c-Fos expression in several brain structures including cingulate and piriform cortex, cortico-amygdaloid and hippocampo-amygdaloid transition zones, hippocampus, hypothalamus, supramammillary nucleus, and centromedial nucleus of thalamus. An intraperitoneal injection of 2 g/kg alcohol prior to stress decreased c-Fos expression in several but not all of these structures. In particular, alcohol strongly attenuated the stress-induced expression of c-Fos in hippocampus and cingulate cortex. Using slot-blot hybridization, significant induction of c-fos mRNA after restraint stress was demonstrated both in hippocampus and cortex, but prior alcohol exposure specifically attenuated c-fos induction only in the hippocampus. The response of c-fos mRNA expression to stress and alcohol differed from the effects on jun-B, c-jun and jun-D mRNA levels. Perhaps surprisingly, acute exposure to alcohol in otherwise unstressed rats did not induce significant changes in expression of IEGs in comparison to control (saline-injected) animals even with doses sufficient to elevate plasma corticosterone. In summary, these studies demonstrate a selective sensitivity of stress-induced activity of neurons of hippocampus and cingulate cortex to acute alcohol exposure.

BACKGROUND

The Burkholderia cepacia complex is associated with colonization or disease in patients with cystic fibrosis (CF). For patients without CF, this complex is poorly understood apart from its presence in occasional point source outbreaks.

OBJECTIVE

To investigate risk factors for B. cepacia bacteremia in hospitalized, intensive care unit patients without CF.

METHODS

We identified patients with 1 or more blood cultures positive for B. cepacia between May 1, 1996, and March 31, 2002, excluding those with CF. Control patients were matched to case patients by ward, duration of hospitalization, and onset date of bacteremia. Matched analyses were used to identify risk factors for B. cepacia bacteremia.

RESULTS

We enrolled 40 patients with B. cepacia bacteremia into the study. No environmental or other point source for B. cepacia complex was identified, although horizontal spread was suspected. Implementation of contact precautions was effective in decreasing the incidence of B. cepacia bacteremia. We selected 119 matched controls. Age, sex, and race were similar between cases and controls. In multivariable analysis, renal failure that required dialysis, recent abdominal surgery, 2 or more bronchoscopic procedures before detection of B. cepacia bacteremia, tracheostomy, and presence of a central line before detection of B. cepacia bacteremia were independently associated with development of B. cepacia bacteremia, whereas presence of a percutaneous feeding tube was associated with a lower risk of disease.

CONCLUSIONS

B. cepacia complex is an important emerging group of nosocomial pathogens in patients with and patients without CF. Nosocomial spread is likely facilitated by cross-transmission, frequent pulmonary procedures, and central venous access. Infection control measures appear useful for limiting the spread of virulent, transmissible clones of B. cepacia complex.

C. difficile infection (CDI) is rarely reported in cystic fibrosis (CF) patients despite frequent hospitalisations and antibiotic usage. Conversely, the prevalence of CDI in inflammatory bowel disease (IBD) has received increased attention. We investigated components of the IgG-specific humoral immune response to C. difficile toxins A and B in patients with C. difficile-associated diarrhoea (CDAD), IBD patients with CDI, CF patients and healthy controls. Serum anti-toxin IgG was determined by ELISA. Circulating antigen-activated B-cells were investigated using Alexa Fluor 488-labelled toxin A and assessed by flow cytometry. Following induction of differentiation of memory B-cells, toxin A- and B-specific antibody secreting cells (ASCs) were quantified using ELISpot. We present the first data showing levels of serum anti-toxin A and B antibodies were significantly higher in patients with CF (without a history of CDI) than in CDAD patients and were stably maintained over time. Notably, the CDAD patients were significantly older than the CF patients. We also show that circulating toxin A-specific memory B-cells (IgD-negative) can be detected in CDAD patients [0.92 (0.09-1.78)%], and were prominent (5.64%, 1.14%) in two CF patients who were asymptomatic carriers of C. difficile. There was correlation between toxin A- and B-specific ASCs, with significantly higher proportions of the latter seen. In some with CDAD, high serum antibody levels were seen to only one of the two toxins. Mucosal secretion of toxin-specific IgG was detected in an additional group of IBD patients with no history of CDI. We conclude that enhanced and stable humoral immune responses to toxins A and B may protect CF and some IBD patients against CDI. The impaired ability to generate strong and/or sustained toxin-specific antibody and memory B-cell responses may increase susceptibility of older patients to CDI and highlight the need to investigate the role of immune senescence in future studies.

Whether immunologic abnormalities correlate with fatigue severity and functional impairment in chronic fatigue syndrome (CFS) was investigated. Blood mononuclear cells were immunophenotyped and circulating ex vivo-produced cytokines were measured in 76 CFS patients and 69 healthy matched controls. Expression of CD11b on CD8 cells was significantly decreased in CFS patients. However, the previously reported increased expression of CD38 and HLA-DR was not confirmed. There was no obvious difference in apoptosis in leukocyte cultures, circulating cytokines, and ex vivo production of interleukin (IL)-1 alpha and IL-1 receptor antagonist. Endotoxin-stimulated ex vivo production of tumor necrosis factor-alpha and IL-beta was significantly lower in CFS. The immunologic test results did not correlate with fatigue severity or psychologic well-being was measured by Checklist Individual Strength, Beck Depression Inventory, and Sickness Impact Profile. Thus, these immunologic tests cannot be used as diagnostic tools in individual CFS patients.

OBJECTIVE

To detect and characterise enterovirus RNA in skeletal muscle from patients with chronic fatigue syndrome (CFS) and to compare efficiency of muscle energy metabolism in enterovirus positive and negative CFS patients.

METHODS

Quadriceps muscle biopsy samples from 48 patients with CFS were processed to detect enterovirus RNA by two stage, reverse transcription, nested polymerase chain reaction (RT-NPCR), using enterovirus group specific primer sets. Direct nucleotide sequencing of PCR products was used to characterise the enterovirus. Controls were 29 subjects with normal muscles. On the day of biopsy, each CFS patient undertook a subanaerobic threshold exercise test (SATET). Venous plasma lactate was measured immediately before and after exercise, and 30 minutes after testing. An abnormal lactate response to exercise (SATET+) was defined as an exercise test in which plasma lactate exceeded the upper 99% confidence limits for normal sedentary controls at two or more time points.

RESULTS

Muscle biopsy samples from 20.8% of the CFS patients were positive for enterovirus sequences by RT-NPCR, while all the 29 control samples were negative; 58.3% of the CFS patients had a SATET+ response. Nine of the 10 enterovirus positive cases were among the 28 SATET+ patients (32.1%), compared with only one (5%) of the 20 SATET- patients. PCR products were most closely related to coxsackie B virus.

CONCLUSIONS

There is an association between abnormal lactate response to exercise, reflecting impaired muscle energy metabolism, and the presence of enterovirus sequences in muscle in a proportion of CFS patients.

BACKGROUND

A unique subset of patients with chronic fatigue syndrome (CFS) and IgM serum antibodies to cytomegalovirus (HCMV) non-structural gene products p52 and CM2 (UL 44 and UL 57) has been described.

METHODS

Fifty-eight CFS patients and 68 non-CFS matched controls were studied. Serum antibodies to EBV viral capsid antigen (VCA) IgM and EBV Early Antigen, diffuse (EA, D) as well HVCMV(V), IgM and IgG; VP (sucrose, density purified V); p52 and CM2 IgM serum antibodies were assayed.

RESULTS

Mean age of CFS patients was 44 years (75% women). Control patients were 9 years older (73% women). Serum EBV VCA IgM positive antibody titers were identified in 33 CFS patients (Group A subset EBV VCA IgM 62.3+/-8.3, neg. <20), but were not present in other CFS patients, (Group B subset EBV VCA IgM 6.8+/-0.7) controls (p<0.0001). EBV VCA IgM titers remained positive in CFS patients from Group A for 24-42 months.

CONCLUSIONS

Serum antibody to EBV VCA IgM may be a specific diagnostic test for a second subset of CFS patients.

OBJECTIVE

Chronic fatigue syndrome (CFS) and fibromyalgia (FM) are medically unexplained conditions that often have overlapping symptoms, including sleep-related complaints. However, differences between the 2 conditions have been reported, and we hypothesized that dynamic aspects of sleep would be different in the 2 groups of patients.

METHODS

Subjects were 26 healthy control subjects, 14 patients with CFS but without FM (CFS alone), and 12 patients with CFS and FM (CFS+FM)-all women.

RESULTS

We studied transition probabilities and rates between sleep stages (waking, rapid eye movement [REM] sleep, stage 1 [S1], stage 2 [S2], and slow-wave sleep [SWS]) and duration distributions of each sleep stage. We found that the probability of transition from REM sleep to waking was significantly greater in subjects with CFS alone than in control subjects, which may be the specific sleep problem for people with CFS alone. Probabilities of (a) transitions from waking, REM sleep, and S1 to S2 and (b) those from SWS to waking and S1 were significantly greater in subjects with CFS+FM than in control subjects; in addition, rates of these transitions were also significantly increased in subjects with CFS+FM. Result (a) might indicate increased sleep pressure in subjects with CFS+FM whereas result (b) may be the specific sleep problem of subjects with CFS+FM. We also found that shorter durations of S2 sleep are specific to patients with CFS+FM, not to CFS alone.

CONCLUSIONS

These results suggest that CFS and FM may be different illnesses associated with different problems of sleep regulation.

The role of CFTR deficiency in promoting inflammation remains unclear. Perez et al. [A. Perez, A.C. Issler, C.U. Cotton, T.J. Kelley, A.S. Verkman and P.B. Davis, CFTR inhibition mimics the cystic fibrosis inflammatory profile. Am J Physiol Lung Cell Mol Physiol 2007; 292:L383-L395.] recently demonstrated that the inhibition of function of w/t CFTR produces an inflammatory profile that resembles that observed in CF patients, whereas we found that correction of F508del-CFTR function with MPB-07 down-modulates the inflammatory response to P. aeruginosa in CF bronchial cells [M.C. Dechecchi, E. Nicolis, V. Bezzerri, A. Vella, M. Colombatti, B.M. Assael, et al., MPB-07 reduces the inflammatory response to Pseudomonas aeruginosa in cystic fibrosis bronchial cells. Am J Respir Cell Mol Biol 2007; 36, 615-624.]. Since both evidence support a link between CFTR function and inflammation, we extended our investigation to other F508del-CFTR correctors, such as miglustat (Norez, 2006), an approved drug for Gaucher disease, in comparison with the galactose analogue NB-DGJ. We report here that miglustat but not NB-DGJ restores F508del-CFTR function in CF bronchial epithelial IB3-1 and CuFi-1 cells. Miglustat and NB-DGJ reduce the inflammatory response to P. aeruginosa in both CF and non-CF bronchial cells, indicating that the anti-inflammatory effect is independent of the correction of F508del-CFTR function. Miglustat also inhibits the inflammatory response induced by the supernatant of mucopurulent material obtained from the lower airway tract of cystic fibrosis patients with chronic bacterial colonization (Ribeiro, 2005). Both compounds do not interfere with the adherence of P. aeruginosa to the cells and reduce the expression of IL-8 not only after challenge with P. aeruginosa but also after exposure to TNF alpha or IL-1 beta, suggesting an effect on transduction proteins downstream and in common with different receptors for pathogens. Finally, miglustat has no major effects on overall binding activity of transcription factors NF-kappaBNF-kB and AP-1. Since miglustat is an approved drug, it could be investigated as a novel anti-inflammatory molecule to ameliorate lung inflammation in CF patients.