Efficacy and Long-Term Outcomes of Palivizumab Prophylaxis to Prevent Respiratory Syncytial Virus Infection in Infants with Cystic Fibrosis in Northern Ireland

BACKGROUND: RSV causes considerable morbidity and mortality in children. In cystic fibrosis (CF) viral infections are associated with worsening respiratory symptoms and bacterial colonization. Palivizumab is effective in reducing RSV hospitalization in high risk patient groups. Evidence regarding its effectiveness and safety in CF is inconclusive. CF screening in N. Ireland enabled timely palivizumab prophylaxis, becoming routine in 2002.

OBJECTIVES: To determine the effect of palivizumab on RSV-related hospitalization and compare lung function and bacterial colonization at age 6 years for those born pre- and post-introduction of palivizumab prophylaxis.

METHODS: A retrospective audit was conducted for all patients diagnosed with CF during the period from 1997 to 2007 inclusive. RSV-related hospitalization, time to Pseudomonas aeruginosa (PA) 1st isolate, lung function and growth parameters were recorded. Comparisons were made for outcomes pre- and post-introduction of routine palivizumab administration in 2002. A cost evaluation was also performed.

RESULTS: Ninety-two children were included; 47 pre- and 45 post-palivizumab introduction. The overall RSV-positive hospitalization rate was 13%. The relative risk of RSV infection in palivizumab non-recipients versus recipients was 4.78 (95%CI: 1.1-20.7), P = 0.027. Notably, PA 1st isolate was significantly earlier in the palivizumab recipient cohort versus non-recipient cohort (median 57 vs. 96 months, P < 0.025) with a relative risk of 2.5. Chronic PA infection at 6 years remained low in both groups, with similar lung function and growth parameters. Total costs were calculated at £96,127 ($151,880) for the non-recipient cohort versus £137,954 ($217,967) for the recipient cohort.

CONCLUSION: Palivizumab was effective in reducing RSV-related hospitalization infection in CF patients. Surprisingly, we found a significantly earlier time to 1st isolate of PA in palivizumab recipients which we could not explain by altered or improved diagnostic tests. 

Production of extended-spectrum β-lactamases and the potential indirect pathogenic role of Prevotella isolates from the cystic fibrosis respiratory microbiota

Extended-spectrum β-lactamase (ESBL) production and the prevalence of the β-lactamase-encoding gene blaTEM were determined in Prevotella isolates (n=50) cultured from the respiratory tract of adults and young people with cystic fibrosis (CF). Time-kill studies were used to investigate the concept of passive antibiotic resistance and to ascertain whether a β-lactamase-positive Prevotella isolate can protect a recognised CF pathogen from the action of ceftazidime in vitro. The results indicated that approximately three-quarters (38/50; 76%) of Prevotella isolates produced ESBLs. Isolates positive for ESBL production had higher minimum inhibitory concentrations (MICs) of β-lactam antibiotics compared with isolates negative for production of ESBLs (P<0.001). The blaTEM gene was detected more frequently in CF Prevotella isolates from paediatric patients compared with isolates from adults (P=0.002), with sequence analysis demonstrating that 21/22 (95%) partial blaTEM genes detected were identical to blaTEM-116. Furthermore, a β-lactamase-positive Prevotella isolate protected Pseudomonas aeruginosa from the antimicrobial effects of ceftazidime (P=0.03). Prevotella isolated from the CF respiratory microbiota produce ESBLs and may influence the pathogenesis of chronic lung infection via indirect methods, including shielding recognised pathogens from the action of ceftazidime.

Lipopolysaccharide modification in Gram-negative bacteria during chronic infection

The Gram-negative bacterial lipopolysaccharide (LPS) is a major component of the outer membrane that plays a key role in host-pathogen interactions with the innate immune system. During infection, bacteria are exposed to a host environment that is typically dominated by inflammatory cells and soluble factors, including antibiotics, which provide cues about regulation of gene expression. Bacterial adaptive changes including modulation of LPS synthesis and structure are a conserved theme in infections, irrespective of the type or bacteria or the site of infection. In general, these changes result in immune system evasion, persisting inflammation, and increased antimicrobial resistance. Here, we review the modifications of LPS structure and biosynthetic pathways that occur upon adaptation of model opportunistic pathogens (Pseudomonas aeruginosa, Burkholderia cepacia complex bacteria, Helicobacter pylori and Salmonella enterica) to chronic infection in respiratory and gastrointestinal sites. We also discuss the molecular mechanisms of these variations and their role in the host-pathogen interaction.

Inhibition of protease-ENaC signaling improves mucociliary function in cystic fibrosis airways

Rationale: In cystic fibrosis (CF) a reduction in airway surface liquid (ASL) height
compromises mucociliary clearance, favoring mucus plugging and chronic bacterial infection. Inhibitors of ENaC have therapeutic potential in CF airways to reduce the hyperstimulated sodium and fluid absorption to levels which can restore airways hydration.

Objectives: To determine whether a novel compound (QUB-TL1) designed to inhibit protease/ENaC signaling in CF airways restores ASL volume and mucociliary function.

Methods: Protease activity was measured using fluorogenic activity assays. Differentiated primary airway epithelial cell cultures (F508del homozygotes) were used to determined ENaC activity (Ussing chamber recordings), ASL height (confocal microscopy) and mucociliary function (by tracking the surface flow of apically applied microbeads). Cell toxicity was measured by LDH assay.

Measurements and Results: QUB-TL1 inhibits extracellularly-located CAPs, including prostasin, matriptase and furin, the activities of which are observed at excessive levels at the apical surface of CF airway epithelial cells (AECs). QUB-TL1-mediated CAPs inhibition results in diminished ENaC-mediated Na+ absorption in CF AECs due to internalization of a prominent pool of cleaved (active) ENaCγ from the cell surface. Importantly, diminished ENaC activity correlates with improved airway hydration status and mucociliary clearance. We further demonstrate QUB-TL1-mediated furin inhibition, which is in contrast to other serine protease inhibitors (camostat mesylate and aprotinin), affords protection against neutrophil elastase-mediated ENaC activation and Pseudomonas aeruginosa exotoxin A induced cell death.

Conclusions: QUB-TL1 corrects aberrant CAP activities providing a mechanism to delay or prevent the development of CF lung disease in a manner independent of CFTR mutation.

Elective versus symptomatic antibiotic treatment in cystic fibrosis patients with chronic Pseudomonas infection of the lungs

BACKGROUND:
A previous retrospective study suggested that a policy of regular anti-pseudomonal antibiotic treatment improved pulmonary function and increased survival in patients with cystic fibrosis chronically infected with Pseudomonas species. The results of a prospective multicentre study to compare the effects on pulmonary function and mortality of three monthly elective anti-pseudomonal antibiotic treatment with conventional symptomatic treatment are reported.

METHODS:
Sixty patients with cystic fibrosis, chronically infected with P aeruginosa, were randomised to the two treatment arms (elective or symptomatic) and followed clinically at yearly reviews. The major end points were changes in forced expiratory volume in one second (FEV(1)) and forced vital capacity (FVC). Survival was a secondary end point.

RESULTS:
Patients in the symptomatic group received a mean of three antibiotic treatments each year and those in the elective group received four antibiotic treatments during each year of the study. No significant differences in FEV(1) and FVC were found between the two groups after three years. There was a statistically non-significant higher rate of deaths in the elective group (n = 4), three of which were associated with B cepacia infection, compared with the symptomatic group (n = 0).

CONCLUSIONS:
This study did not demonstrate an advantage of a policy of elective antibiotic treatment over symptomatic treatment in patients with cystic fibrosis chronically infected with Pseudomonas species.

Enantiocomplementary preparation of (S)- and (R)-1-pyridylalkanols via ketone reduction and alkane hydroxylation using whole cells of Pseudomonas putida UV4

A previously unreported alcohol dehydrogenase enzyme in the mutant soil bacterium Pseudomonas putida UV4 catalyses the reduction of 2-, 3- and 4-acylpyridines to afford the corresponding (S)-1-pyridyl alkanols, with moderate to high e.e., whilst under the same conditions 2,6-diacetylpyridine is readily converted to the corresponding enantiopure C2-symmetric (S,S)-diol in one step. In contrast, the toluene dioxygenase enzyme in the same organism catalyses the hydroxylation of 2- and 3-alkylpyridines to (R)-1-(2-pyridyl) and (R)-1-(3-pyridyl)alkanols. This combination of oxidative and reductive biotransformations thus provides a method for preparing both enantiomers of chiral 1-pyridyl alkanols using one biocatalyst.

The effect of polyphosphate kinase gene deletion on polyhydroxyalkanoate accumulation and carbon metabolism in Pseudomonas putida KT2440.

The primary enzyme involved in polyphosphate (polyP) synthesis, polyP kinase (ppk), has been deleted in Pseudomonas putida KT2440. This has resulted in a threefold to sixfold reduction in polyhydroxyalkanoate (PHA) accumulation compared with the wild type under conditions of nitrogen limitation, with either temperature or oxidative (H2O2) stress, when grown on glucose. The accumulation of PHA by Δppk mutant was the same as the wild type under nitrogen-limiting growth conditions. There was no difference in polyP levels between wild-type and Δppk strains under all growth conditions tested. In the Δppk mutant proteome, polyP kinase (PPK) was undetectable, but up-regulation of the polyp-associated proteins polyP adenosine triphosphate (ATP)/nicotinamide adenine dinucleotide (NAD) kinase (PpnK), a putative polyP adenosine monophosphate (AMP) phosphotransferase (PP_1752), and exopolyphosphatase was observed. Δppk strain exhibited significantly retarded growth with glycerol as carbon and energy source (42 h of lag period compared with 24 h in wild-type strain) but similar growth to the wild-type strain with glucose. Analysis of gene transcription revealed downregulation of glycerol kinase and the glycerol facilitator respectively. Glycerol kinase protein expression was also downregulated in the Δppk mutant. The deletion of ppk did not affect motility but reduced biofilm formation. Thus, the knockout of the ppk gene has resulted in a number of phenotypic changes to the mutant without affecting polyP accumulation.