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Modulation of mucosal immune responses to pneumococcal protein antigens in
human NALT by TLR2 and TLR9 ligands
Qibo Zhang1, Linda Bagrade2, Ed Clark1, James Paton4, Desmond Nunez 3, Adam Finn 1,2
Departments of 1Cellular & Molecular Medicine, 2Clinical Science at South Bristol and
3Clinical Science at North Bristol, University of Bristol, 4School of Molecular and Biomedical
Science, University of Adelaide, Australia.
Background:
The ability of TLR ligands to activate immune cells has been regarded as the basis for
adjuvant activity. Bacterial lipopeptide (BLP) and CpG-DNA which are known as TLR2 and
TLR9 ligands respectively, have been suggested to be candidate mucosal adjuvants. We
investigated whether BLP and CpG-DNA enhance mucosal B cell antibody production to
pneumococcal protein antigens in adenoids which are part of nasal-associated lymphoid
tissue (NALT) in children.
Methods:
Adenoidal mononuclear cells (MNC) were isolated followed by B and T cell separation
using MACS. Memory and naïve T or B cells were separated by either CD45RO+, or
CD27+ cell depletion. MNC or combinations of memory T and B Cells or naïve T and B
cells were cultured with pneumococcal protein antigens, with BLP or CpG-DNA. Antibody
production was measured by immunoassay and cell proliferation was assessed by CFSE
assay. Expressions of costimulatory molecules including B7.1/7.2, B7h, and CD28 and
inducible costimulatory molecule (ICOS)) were analysed by flow-cytometry.
Results:
Co-stimulation with BLP or CpG-DNA significantly enhanced the primary IgG and IgM
antibody responses to CbpA and PspA (p<0.01). While CpG-DNA also increased memory
IgG responses to the antigens, BLP reduced them. BLP also enhanced primary and
suppressed memory CD4 T cell proliferation, induced significant upregulation of B7h on
macrophage/monocytes and its ligand ICOS on memory CD4 T cells with associated IL10
production. Anti-TLR2 antibody inhibited BLP-induced primary T and B cell responses. AntiTLR2, -B7h and -IL10 antibodies blocked suppression of memory T and B cell responses
by BLP.
Conclusion:
Both BLP and CpG-DNA can enhance primary, but differentially modulate memory-type
mucosal CD4 T cell and B cell responses to pneumococcal protein antigens in human
NALT. These effects of BLP are probably via TLR2 are modulated by costimulatory
molecules. This may have important implications for vaccination strategies against
pneumococcus.
Persistence of protection with conjugate vaccines
Andrew J Pollard.
Oxford Vaccine Group, Department of Paediatrics, University of Oxford
Polysaccharide-encapsulated organisms, such as Haemophilus influenzae type b,
Streptococcus pneumoniae and Neisseria meningitidis, are the leading cause of the
serious bacterial diseases of childhood including meningitis and pneumonia and the
predominant cause of death among children less than 5 years of age in the world.
Vaccines based on the polysaccharide capsule are poorly immunogenic in early
childhood, presumably because of immaturity of the splenic marginal zone, and as T independent antigens, do not induce immunological memory at any age and may even
interfere with later immune responses. The development and implementation of T dependent protein-polysaccharide conjugate vaccines over the past 2 decades has
dramatically reduced the burden of disease and mortality from these organisms
wherever they have been used. After immunisation, in addition to direct protection of
the immunised, herd immunity is induced further decreasing disease amongst those
who remain susceptible including the non- and incompletely immunised. In the United
Kingdom, Group C meningococcal (MenC) vaccine was introduced in 1999 with a
massive catch up campaign in which all individuals under the age of 19 (later
extended to 24) years were immunised with the conjugate. MenC disease fell amongst
both the immunised and unimmunised sections of the population from almost 1000
cases in 1999 to only 28 in 2006. Significantly, the antibody produced after infant
immunization, even with 3 doses of MenC vaccine does not persist well and the levels
will have fallen below the protective threshold in 50% of infants by a year of age and
as few as 12% have persistent seroprotection by 4 years of age. Persistence is better
later in childhood. Herd immunity hides the rapid waning of vaccine-induced immunity
among young children and booster doses of vaccine were implemented in 2006 in the
second year of life to sustain protection. We have extrapolated data to provide
information on the current seroprotection rates in the UK population . At the end of
2008 the vast majority of children aged 3-14 years are susceptible to group C
meningococcal disease, but are presumably protected through herd immunity as
disease is currently so rare. However, over the next decade, these children will age to
13-24 years, the cohort with historically high rates of disease and MenC may resurge.
Sustained population immunity for MenC might be best achieved over the decades to
come by the addition of an adolescent booster of MenC or MenACYW vaccine.
Title: Structure and Function of Streptococcus agalactiae Esat-6
A Shukla, N Attwood, R Whitehead, D Biswas, R Shaw, K Lightbody, P Renshaw, M Carr,
L Snyder, R May, M Neely, T Lammas, M Pallen, S White, Mark Anthony.
Birmingham Women's Hospital & The School of Biosciences, University of Birmingham
Aims: To explore the significance of an esat-6 homologue in S. agalactiae (group B
streptococcus).
Methods: An esat-6 homologue was identified in 7 of 8 S. agalactiae genomes. The
gene was mutated, and the virulence of mutants assessed in a zebrafish model, and
their survival examined in a human macrophage killing assay. Recombinant S.
agalactiae Esat-6 was produced for crystal structure determination. We examined
secretion of Esat-6 by Western blot, and fluorophore-tagged rEsat-6 to assess binding
to human macrophages. rEsat-6 from M. tuberculosis, S. aureus and S. agalactiae,
and synthetic peptides representing their C-termini, were assessed for their ability to
inhibit Toll-Like-Receptors in mouse and human macrophages, stimulated with TLR2,
3 and 4 ligands.
Results: We solved the crystal structure of Esat-6 from Streptococcus agalactiae (see Fig).
S. agalactiae Esat-6 has a helix-turn-helix homodimer configuration and has the capacity to
form polymeric fibrillar strands with overlapping C-termini. We show that Esat-6 is secreted
by S. agalactiae, that it is critical for virulence, that it binds to macrophages, and impedes
TLR2 via its C-terminus. We also show that Staphylococcus aureus Esat-6 impedes TLR2,
probably through interaction with different aspects of the extra cellular domain of TLR2.
Conclusions: The discovery that S. aureus and S. agalactiae Esat-6 family proteins, each
with different C-terminal amino acid compositions, impede TLR2 is unexpected. The
structural, virulence and TLR2 findings enhance our understanding of Esat-6-family
proteins and of the pathogenesis of S. agalactiae and S. aureus.
Interferon- gamma release assays do not identify more children with active TB
than TST
Beate Kampmann 1,2,4, Elizabeth Whittaker 1, Amanda Williams 5, Sam Walters 3, Andrea
Gordon 2, Nuria Martinez-Alier1,3, Bhanu Williams 5, Angela M Crook 6, Anne-Marie
Hutton 2, Suzanne T Anderson 1,7
Affiliations: 1 Academic Department of Paediatric Infectious Diseases
Imperial College London. 2 Wellcome Centre for Clinical Tropical Medicine, Imperial College
London. 3 Imperial College NHS Trust, St. Mary’s Campus, London. 4 Centre for Respiratory
Infection, Imperial College London. 5 Department of Paediatrics, Northwest London Hospital
Trusts, Northwick Park, London. 6 MRC Clinical Trials Unit, MRC, Euston Road, London.
7
Brighton and Sussex Medical School, University of Sussex, Falmer, Brighton BN1 9PS, UK
Rationale:
The diagnosis of tuberculosis remains a challenge in children. Lately, blood based
assays that measure the release of interferon-gamma in responses to TB-specific
antigens (IGRA) have been developed and were introduced into the NICE guidelines
for management of TB in the UK in 2006. Although primarily recommended for
screening for latent TB infection (LTBI), many clinicians also wish to employ IGRA as
a diagnostic test for active tuberculosis (TB).
Data in children being investigated for TB using either IGRA are sparse, and no sideby-side comparison of the commercially available assays and the TST has been
published from the UK.
Objective:
We compared the performance of the two commercially available IGRA and tuberculin
skin test (TST) side-by-side in children with active TB and LTBI.
Methods:
We conducted a prospective study of 209 children investigated for active (n=91) or
latent tuberculosis (n=118). We simultaneously used TST, Quantiferon -Gold–in tube
(QFG-IT) and T-Spot.TB assays.
Results:
For culture- confirmed active TB (n=25), the sensitivity of the TST > 15 mm was 83%,
compared to 80% for QFG-IT and 58% for T-Spot.TB. IGRA did not perform
significantly better than TST, although QFG-IT was significantly better than T-Spot.TB
(p=0.012). The agreement between QFG-IT and T-Spot.TB in culture-confirmed TB
was poor at 66.7% (κ 0.15).
Conclusions:
A negative IGRA should not dissuade paediatricians from diagnosing and treating
presumed active TB. If used for diagnosis of LTBI, IGRA could significantly reduce the
numbers of children receiving chemoprophylaxis with very good concordance between
both tests.
Funding: Wellcome Trust (BK), European Society for Paediatric Infectious Diseases, PEEL Foundation.
Supported by the Biomedial Research Centre (BRC) at Imperial College
Effects of prenatal probiotic treatment on infant immune responses and colonisation
patterns
1, 2 Robert
J Boyle MB ChB, 2Lahtinen S PhD, 1Mah L-J BSc, 1Kivuori S MSc, 1Chen A
MD, 2Robins-Browne RM PhD, 1, 2Mimi L-K Tang MBBS PhD
1 Murdoch Children’s Research Institute; 2University of Melbourne, Victoria, Australia
Aims:
Observational studies suggest that microbial exposures may be important in
preventing allergic immune responses. Intervention trials have identified perinatal
administration of probiotic bacteria such as Lactobacillus rhamnosus GG (LGG) as a
promising approach for preventing allergic disease. We investigated the mechanisms
of action through which LGG may prevent allergic disease. We evaluated the effects of
LGG administration to pregnant women on infant immune responses and on infant
intestinal microbiota composition.
Methods:.
In a randomised controlled trial of LGG treatment during pregnancy cord blood
mononuclear cells from 73 participants were evaluated for markers of T cell regulation,
antigen presenting cell phenotype, cytokine secretion and proliferative response to a
range of in vitro stimuli. Proliferative response, markers of T cell regulation and
antigen presenting cell phenotype were also assessed in peripheral blood
mononuclear cells from 11 healthy adults before and after LGG treatment. Rectal and
vaginal swabs, faeces and breast milk samples from participants or their infants were
evaluated for the presence of LGG by culture and strain-specific PCR. Bifidobacterium
spp. composition was evaluated in rectal swabs and faeces samples using terminal
restriction fragment length polymorphism.
Results:
In healthy adults LGG treatment was associated with a 30% reduction in T cell
proliferation to heat killed LGG (P=0.03). Treatment of pregnant women with LGG was
not associated with any change in cord blood mononuclear cell immune responses.
Treatment of pregnant women increased LGG detection in maternal rectal swabs
taken at birth, but did not influence infant colonisation with LGG during the first 90
days. Prenatal LGG treatment increased B. longum colonisation in infants at 90 days
(present in 82% probiotic group, 61% placebo group; P=0.01). In infants whose
mothers received prenatal LGG the Bifidobacterium microbiota more closely
resembled that of healthy breast-fed infants.
Conclusions:
Prenatal treatment with the probiotic LGG does not induce immune tolerance or
priming by trans-placental signalling to the human foetus. Treatment may however
influence the infant intestine by promoting a healthy Bifidobacterium microbiota.
STAT3 Activation in Paediatric Solid Tumours
Fyeza Hasan 1,a, Sian Gibson 2, Dyanne Rampling 2 Neil Sebire 2, Oliver Campos 2, Tom
Jacques 2, Adrienne Flanagan 3 and John Anderson 1. Molecular Haematology and
Cancer Biology Unit, UCL Institute of Child Health, London 1, Histopathology Unit,
Great Ormond Street Hospital, London 2, Institute of Orthopaedics and Musculoskeletal
Science, UCL 3. Funded by Institute of Child Health and Great Ormond Street Hospital
Biomedical Research Centre Grant a.
Aims:
Cancers use many different mechanisms to avoid immune detection. Activating STAT3
(signal transducer and activator of transcription 3) is one such mechanism. There is
evidence that STAT3 is phosphorylated and activated in a large number of adult
cancers, allowing progression by stimulating growth and also by inhibiting the immune
response to cancer. Furthermore, STAT3 activation may limit the effects of
immunotherapeutic treatments for cancer. This study investigated whether STAT3 is
activated in paediatric solid tumours.
Methods:
Tissue arrays were produced using archived tumour samples for 9 types of paediatric
solid tumour, and immunohistochemistry was performed on these arrays to look for
nuclear staining for phosphorylated STAT3. Staining was quantified us ing Image J
image analysis software. Consecutive 4mm sections from tissue arrays were also
stained for the T-cell, T-regulatory cell and macrophage markers, CD3, FOXP3 and
CD68, and correlations made with phosphorylated STAT3 staining patterns.
Results:
Approximately half of the Wilms’ tumours and Ewing’s sarcomas demonstrated STAT3
activation. A lower proportion of other tumours including medulloblastomas,
ependymomas, rhabdoid tumours, neuroblastomas and osteosarcomas were STAT3
activated. Staining for immune cell markers demonstrated that the majority of STAT3
phosphorylated cells were unlikely to be tumour infiltrating T-cells or T-regulatory cells.
A number of tumour types demonstrated macrophage infiltration.
Conclusions:
STAT3 may be an appropriate target in a subset of paediatric cancers. It is possible
that STAT3 activation contributes to the immune inhibition seen within the
microenvironment of these tumours. This work is part of a Cancer Research UK
funded collaboration with the UCL School of Pharmacy to develop small molecule
inhibitors of STAT3 and it is hoped that inhibitors developed during this project will be
synergistic with immunotherapeutic strategies.
A dyad of Lymphoblast Cysteine Proteases Degrades the Key Anti -Leukaemic
Drug L-Asparaginase
Naina Patel1, Shekhar Krishnan 1, Marc Offman 2, Marcin Krol 2, Catherine Moss 3, Carly
Leighton 1, Frederik van Delft 1, Hany Ariffin 4, 1Mark Holland, 1Jizhong Liu, 1Seema
Alexander, 1Clare Dempsey, 1Ashish Masurekar, Monica Essink 5, Colin Watts 3, Paul
Bates 2, Vaskar Saha 1
1 CRUK Children’s Cancer Group, Paterson Institute for Cancer Research, Manchester,
2 Biomolecular Modelling Laboratory, CRUK London Research Institute, London,
3 Division of Cell Biology and Immunology, School of Life Sciences, Dunde e,
4 Department of Paediatrics, University Malaya, Malaysia, 5 Medac GmbH, Germany
L-Asparaginase (Asnase) is a key drug in the therapy of childhood acute lymphoblastic
leukaemia (ALL). Drug resistance commonly occurs through development of
hypersensitivity and formation of neutralising antibodies. We have identified two
lysosomal cysteine proteases in B-lineage lymphoblasts, Cathepsin B (CTSB) and
Asparaginyl Endopeptidase (AEP) that specifically degrade and inactivate Asnase.
Gene expression analysis, verified by protein studies in a selected number, show that
all leukaemic cells express CTSB whereas AEP is predominantly expressed in poor
risk cytogenetic subtypes of B-lineage ALL. AEP cleavage sites were identified and
the known crystallographic structure of Asnase was used to predict the molecular
consequences of AEP cleavage. These predictions were subsequently validated using
site-directed mutageneses and biochemical analyses. AEP cleaves Asnase
sequentially from the N-terminus and inactivates the molecule without disrupting
known antigenic epitopes. Modifying the first cleavage site generates an enzymatically
active Asnase variant resistant to AEP cleavage - such a compound would have
therapeutic potential.
We speculate that Asnase degradation is basally mediated by CTSB but is augmented
by AEP. Thus screening for AEP prior to treatment may permit further optimisation of
therapy with Asnase. Our investigations have identified a hitherto unknown pathway
for Asnase degradation and inactivation and suggest a novel mechanism of drug
resistance in childhood ALL and a model for the development of a new generation of
more effective asparaginases
Confocal Endomicroscopy: A New Tool in the in vivo Diagnosis of Coeliac
Disease
Krishnappa Venkatesh 1, Ashraf Abou-Taleb 1 Marta Cohen 2, Clair Evans 2, Christopher
Taylor1, Mike Thomson 1.
1 Centre for Paediatric Gastroenterology, Sheffield
Children’s NHS Foundation Trust, Sheffield, United Kingdom. 2 Department of
Histopathology, Sheffield Children’s NHS Foundation Trust, Sheffield, United Kingdom
Background and aims
Confocal laser endomicroscopy (CLE) is a recent development which enables surface
and subsurface imaging of living cells in vivo at x1000 magnification. The aims of the
present study were to define confocal features of coeliac disease and to evaluate the
usefulness of the CLE in the diagnosis of coeliac disease in children in comparison to
histology.
Methods
9 patients (7 female) with a median age 8.1 years (range 2-10.5) and weight of 23 kg
(range 10.5 -71) with positive coeliac serology and 10 matched controls underwent
oesophago-gastro-duodenoscopy (OGD) using the confocal laser endomicroscope
(EC3870CILK; Pentax, Tokyo, Japan). Intravenous sodium fluorescein and topical
acriflavine were used as contrast agents. Histology of coeliac disease was graded
according to Marsh classification. Confocal features of coeliac disease were defined
prior to blinding. These included loss of surface villous architecture, presence of broad
villi, infolding of villi, intervillous bridging (“sticky villi”) and decreased goblet cells in
Marsh type 3b (partial villous atrophy) and absence of villi, crypt hypertrophy and
decreased goblet cells in Marsh type 3c (total villous atrophy). Histologic sections
were compared with same site confocal images by 2 experienced paediatric
histopathologists and endoscopists, who were blinded to the diagnosis, respectively.
Results
The median procedure time for OGD was 16.4 minutes (range 8-25). A total of 1273
confocal images from both patients and controls were compared with 44 same site
duodenal biopsies. 6 patients with coeliac disease had crypt hypertrophy and total
villous atrophy (Marsh type 3c) and 3 had crypt hypertrophy with partial villous atrophy
(Marsh type 3b). Sensitivity and specificity for the diagnosis of coeliac disease were
100% and 89% with a kappa coefficient for inter-observer agreement between the
paediatric gastroenterologists was 0.758. In addition 74% of the images were
considered to be of good quality.
Conclusion
Confocal endomicroscopy offers the prospect of diagnosis of coeliac disease during
ongoing endoscopy. It also enables targeting biopsies to abnormal mucosa and
thereby increasing the diagnostic yield especially when villous atrophy is patchy in the
duodenum.
Fat or glucose? The effect of elective caesarean section on the end product of
hepatic glycerol metabolism 7 days post-partum.
Matthew J. Hyde 1, Julian L. Griffin 2, Neena Modi 1, Lynne Clarke 3 and Paul R. Kemp 4
1
Neonatal Medicine, Imperial College London, Chelsea and Westminster Hospital, 369
Fulham Road, London, SW10 9NH, UK; 2Department of Biochemistry, University of
Cambridge, Hopkins Building, Tennis Court Road, Cambridge, CB2 1QW, UK; 3School of
Agriculture, Policy & Development, University of Reading, RG6 6AR, UK; 4Section of
Molecular Medicine, Sir Alexander Fleming Building, Imperial College London, South
Kensington, SW7 2AZ, UK.
Caesarean section (CS) delivered piglets accumulate less hepatic lipid after 7 days on total
parenteral nutrition (TPN), than vaginally delivered (VD) animals (1) . Babies born by elective CS
have altered plasma hormone and metabolite profiles compared to VD neonates (2) , including
thyroid hormone (3) . Glycerolphosphate dehydrogenase (G3PDH) activity postpartum is stimulated
by thyroid hormone secretion (4,5) .
We hypothesise that CS neonates fail to stimulate
gluconeogenic activity from glycerol, instead diverting glycerol into hepatic lipid storage.
Method: Experiments were conducted under Home Office licence with relevant ethical approval.
Piglets born by CS (≈3 days preterm) or VD (at term), had bilateral jugular catheters inserted 3
hours postpartum. Piglets (CS n = 5, VD n = 4) received TPN (including Intralipid 20%) for ≈7
days, after which they were killed and tissue sampled.
Hepatic lipid was determined
gravimetrically. Plasma hormone and metabolite concentrations and liver glycerol were assayed
using commercial kits. Liver G3PDH and phosphoenolpyruvate carboxykinase (PEPCK) activity
were measured by colourimetric assay and hepatic glucose by 1H NMR spectroscopy. Statistical
differences were assessed by ANOVA General Linear Model; results are expressed as mean ±
SEM.
Results: Plasma thyroxine (T 4) and triiodothyronine (T 3 ) concentrations were elevated in VD
animals at birth (T 4 : CS, 6.75±0.45; VD 10.09±3.72 nmol∙L -1 , p<0.05. T 3 : CS, 1.43±0.09;
VD 3.75±1.78 nmol∙L -1 , p<0.01) but there was no difference in plasma T 3 on day 7 between CS
and VD piglets.
After 7 days, liver lipid content was higher (p<0.05) in CS vs. VD piglets (CS 7.6±0.8;
VD 4.2±0.2 % (w/w)). Conversely, hepatic and plasma glucose were elevated in the VD piglets
(Hepatic glucose: CS, 8.44±0.27; VD 16.30±0.20 arbitrary units p<0.05. Plasma glucose: CS,
3.80±0.16; VD 5.13±0.76 mmol∙L -1 p=0.17) suggesting greater gluconeogenesis in VD animals.
Hepatic glycerol was reduced (CS 39.8±8.2; VD 14.6±8.4 mg∙g liver -1 : p<0.05) whilst hepatic
G3PDH activity increased (CS 12.27±1.77; VD 23.89±5.81 M∙mg protein -1min-1 : p<0.05) in VD vs.
CS animals. However, mode of delivery did not alter PEPCK activity (CS 10.57±1.38; VD
11.35±0.70 mU∙mg protein -1 ). These data suggest increased gluconeogenic activity from glycerol,
but not from tricarboxylic acid cycle precursors, in VD piglets.
Conclusion: In the neonate, VD increases thyroid hormones which are associated with increased
glycerolphosphate dehydrogenase activity. This change in enzyme activity diverts glycerol
towards gluconeogenesis rather than fatty acid esterification and consequent lipid storage. The
absence of these changes in CS neonates may increase susceptibility to abnormal lipid
accumulation.
The authors wish to thank John Laws, Anne Corson, Kate Perkins and Jennie Litten, for their
assistance with animal husbandry. MJH is funded by a BBSRC Studentship.
The culture of primary bronchial epithelial cells from cystic fibrosis lungs
removed at the time of transplantation - A model to study cystic fibrosis lung
disease
Brodlie M1,2, McKean MC 2, Perry J3, Nicholson A 3, Johnson GE 1, Pearson JP 4, Fisher
A 1, Corris PA 1, Lordan JL 1 and Ward C 1
1
Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, NE2 4HH
Paediatric Respiratory Unit, Newcastle Hospitals NHS Foundation Trust, Freeman Hospital,
Newcastle upon Tyne, NE7 7DN
3
Department of Microbiology, Newcastle Hospitals NHS Foundation Trust
4
Institute for Cell and Molecular Biosciences, Newcastle University
2
Aims
Mortality and morbidity in cystic fibrosis (CF) are largely due to lung disease. Despite
improvements in survival the exact pathogenesis of CF lung disease remains poorly
understood. Ultimately, however, it results in progressive bronchiectasis and premature
death. Lung transplantation is the only life-sustaining option for end-stage disease. Studies
involving animal models and immortalised cell lines have contributed significantly to our
current knowledge of CF lung disease. However, there are inherent limitations to both
approaches, including poor replication of lung pathology and failure to reflect in vivo
findings. The opportunity to work on primary tissue from patients with CF is rare. The aim of
this work was to establish an ex vivo culture system for primary bronchial epithelial cells
(PBECs) from the lungs of people with CF removed at the time of transplantation.
Methods
Pieces of segmental bronchus were removed immediately after explantation and treated
with patient-specific antimicrobials and mucolytics to achieve disinfection. PBECs were
harvested and submerged cultures established before transfer to an air-liquid interface
(ALI). Cultures were characterised morphologically and histologically using light and
scanning electron microscopy. Mucus production at ALI was assessed by enzyme-linked
immunosorbent assay and amylase-periodic acid Schiff staining.
Results
PBECs have been successfully cultured from 12 of 18 patients attempted. Mucus
production and tight junction formation has been demonstrated at ALI. The PBECs
remain viable after storage in liquid nitrogen. PBEC cultures failed from 2 patients due
to immediate overgrowth with Burkholderia cepacia complex and in 4 patients initially
successful cultures overgrew with Pseudomonas aeruginosa once antiimicrobials were
withdrawn.
Conclusions
PBEC culture is possible from lungs removed at the time of transplantation from people
with CF. Tailored antimicrobial strategies are practicable and yield a favourable success
rate. This technique represents a valuable resource that provides a cellular model to
elucidate the pathogenic mechanisms in CF lung disease and to investigate potential
therapeutic targets.