Publications by authors named "Paul Rooney"

94 Publications

One threat, different answers: the impact of COVID-19 pandemic on cornea donation and donor selection across Europe.

Br J Ophthalmol 2020 Nov 26. Epub 2020 Nov 26.

University Eye Clinic Maastricht, Maastricht University Medical Centre+, Maastricht, The Netherlands.

Objectives: To assess to which extent the COVID-19 pandemic affected corneal transplantation by virtue of donor selection algorithms in different European countries.

Design: Survey.

Setting: 110 eye banks in 26 European countries.

Participants: 64 eye banks covering 95% of European corneal transplantation activity.

Interventions: A questionnaire listing the number of corneas procured and distributed from February to May 2018-2020 was circulated to eye banks.

Main Outcome Measures: The primary outcome was the number of corneal procurements. Additional outcomes were national algorithms for donor selection, classified according to their stringency (donors with COVID-19 history, suspected for COVID-19, asymptomatic, PCR testing) and the pandemic severity in each country. We calculated Spearman's correlation coefficient to determine, two by two, the relationship between the 3-month decline in eye banking activity (procurement), the stringency of donor selection algorithm and the grading of pandemic severity (cases and deaths). A partial correlation was run to determine the relationship between decline and stringency while controlling for pandemic severity.

Results: Procurements decreased by 38%, 68% and 41%, respectively, in March, April and May 2020 compared with the mean of the previous 2 years, while grafts decreased, respectively, by 28%, 68% and 56% corresponding to 3866 untreated patients in 3 months. Significant disparities between countries and the decrease in activity correlated with stringency in donor selection independent of pandemic severity.

Conclusions: Our data demonstrate significant differences between countries regarding donor screening algorithms based on precautionary principles and, consequently, a decrease in the donor pool, already constrained by a long list of contraindications. Fundamental studies are needed to determine the risk of SARS-CoV-2 transmission by corneal transplantation and guide evidence-based recommendations for donor selection to justify their substantial medical and economic impact.
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http://dx.doi.org/10.1136/bjophthalmol-2020-317938DOI Listing
November 2020

Gastroesophageal device erosion.

Gastrointest Endosc 2021 Jan 18;93(1):260-261. Epub 2020 Jul 18.

Department of Gastroenterology, Belfast City Hospital, Belfast, Antrim, UK.

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http://dx.doi.org/10.1016/j.gie.2020.07.012DOI Listing
January 2021

Co-expression of SARS-CoV-2 entry genes in the superficial adult human conjunctival, limbal and corneal epithelium suggests an additional route of entry via the ocular surface.

Ocul Surf 2021 01 3;19:190-200. Epub 2020 Jun 3.

Biosciences Institute, Faculty of Medical Sciences, Newcastle University, UK. Electronic address:

Purpose: The high infection rate of SARS-CoV-2 necessitates the need for multiple studies identifying the molecular mechanisms that facilitate the viral entry and propagation. Currently the potential extra-respiratory transmission routes of SARS-CoV-2 remain unclear.

Methods: Using single-cell RNA Seq and ATAC-Seq datasets and immunohistochemical analysis, we investigated SARS-CoV-2 tropism in the embryonic, fetal and adult human ocular surface.

Results: The co-expression of ACE2 receptor and entry protease TMPRSS2 was detected in the human adult conjunctival, limbal and corneal epithelium, but not in the embryonic and fetal ocular surface up to 21 post conception weeks. These expression patterns were corroborated by the single cell ATAC-Seq data, which revealed a permissive chromatin in ACE2 and TMPRSS2 loci in the adult conjunctival, limbal and corneal epithelium. Co-expression of ACE2 and TMPRSS2 was strongly detected in the superficial limbal, corneal and conjunctival epithelium, implicating these as target entry cells for SARS-CoV-2 in the ocular surface. Strikingly, we also identified the key pro-inflammatory signals TNF, NFKβ and IFNG as upstream regulators of the transcriptional profile of ACE2TMPRSS2 cells in the superficial conjunctival epithelium, suggesting that SARS-CoV-2 may utilise inflammatory driven upregulation of ACE2 and TMPRSS2 expression to enhance infection in ocular surface.

Conclusions: Together our data indicate that the human ocular surface epithelium provides an additional entry portal for SARS-CoV-2, which may exploit inflammatory driven upregulation of ACE2 and TMPRSS2 entry factors to enhance infection.
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http://dx.doi.org/10.1016/j.jtos.2020.05.013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7267807PMC
January 2021

Single-stage repair of contaminated hernias using a novel antibiotic-impregnated biologic porcine submucosa tissue matrix.

BMC Surg 2020 Mar 30;20(1):58. Epub 2020 Mar 30.

St. Paul's Hospital, 1081 Burrard Street, Vancouver, BC, V6Z 1Y6, Canada.

Background: Single-stage repair of incisional hernias in contaminated fields has a high rate of surgical site infection (30-42%) when biologic grafts are used for repair. In an attempt to decrease this risk, a novel graft incorporating gentamicin into a biologic extracellular matrix derived from porcine small intestine submucosa was developed.

Methods: This prospective, multicenter, single-arm observational study was designed to determine the incidence of surgical site infection following implantation of the device into surgical fields characterized as CDC Class II, III, or IV.

Results: Twenty-four patients were enrolled, with 42% contaminated and 25% dirty surgical fields. After 12 months, 5 patients experienced 6 surgical site infections (21%) with infection involving the graft in 2 patients (8%). No grafts were explanted.

Conclusions: The incorporation of gentamicin into a porcine-derived biologic graft can be achieved with no noted gentamicin toxicity and a low rate of device infection for patients undergoing single-stage repair of ventral hernia in contaminated settings.

Trial Registration: The study was registered March 27, 2015 at www.clinicaltrials.gov as NCT02401334.
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http://dx.doi.org/10.1186/s12893-020-00715-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7106678PMC
March 2020

Surgical management of large abdominal wall fibromatosis during pregnancy.

BMJ Case Rep 2019 Jul 21;12(7). Epub 2019 Jul 21.

Trauma and Orthopaedic Department, Royal Liverpool and Broadgreen University Hospitals NHS Trust, Liverpool, UK.

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http://dx.doi.org/10.1136/bcr-2018-227811DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6663169PMC
July 2019

Volatile organic compounds emitted from faeces as a biomarker for colorectal cancer.

Aliment Pharmacol Ther 2019 04 3;49(8):1005-1012. Epub 2019 Mar 3.

Royal Liverpool and Broadgreen University Hospital Trust, Liverpool, UK.

Background: Colorectal cancer remains a leading cause of mortality and morbidity. The UK Bowel Cancer Screening Programme (BCSP) has demonstrated that detection of colorectal cancer at an earlier stage and identification of advanced pre-malignant adenomas reduces mortality and morbidity.

Aim: To assess the utility of volatile organic compounds as a biomarker for colorectal neoplasia.

Methods: Faeces were collected from symptomatic patients and people participating in the UK BCSP, prior to colonoscopy. Headspace extraction followed by gas chromatography mass spectrometry was performed on faeces to identify volatile organic compounds. Logistic regression modelling and 10-fold cross-validation were used to test potential biomarkers.

Results: One hundred and thirty-seven participants were included (mean age 64 years [range 22-85], 54% were male): 60 had no neoplasia, 56 had adenomatous polyp(s) and 21 had adenocarcinoma. Propan-2-ol was significantly more abundant in the cancer samples (P < 0.0001, q = 0.004) with an area under ROC (AUROC) curve of 0.76. When combined with 3-methylbutanoic acid the AUROC curve was 0.82, sensitivity 87.9% (95% CI 0.87-0.99) and specificity 84.6% (95% CI 0.65-1.0). Logistic regression analysis using the presence/absence of specific volatile organic compounds, identified a three volatile organic compound panel (propan-2-ol, hexan-2-one and ethyl 3-methyl- butanoate) to have an AUROC of 0.73, with a person six times more likely to have cancer if all three volatile organic compounds were present (P < 0.0001).

Conclusions: Volatile organic compound analysis may have a superior diagnostic ability for the identification of colorectal adenocarcinoma, when compared to other faecal biomarkers, including those currently employed in UK. Clinical trial details: National Research Ethics Service Committee South West - Central Bristol (REC reference 14/SW/1162) with R&D approval from University of Liverpool and Broadgreen University Hospital Trust (UoL 001098).
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http://dx.doi.org/10.1111/apt.15140DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6593415PMC
April 2019

Surgical audit: are we not closing the loop?

Int J Health Care Qual Assur 2018 Oct;31(8):966-972

Royal Liverpool University Hospital , Liverpool, UK.

Purpose: A clinical audit is a key component of the clinical governance framework. The rate of audit completion in general surgery has not been investigated. The purpose of this paper is to assess the rates of audit activity and completion and explore the barriers to successful audit completion.

Design/methodology/approach: This was a multi-centre study evaluating current surgical audit practice. A standardised audit proforma was designed. All clinical audits in general surgery during a two-year period were identified and retrospectively reviewed. Data held by the audit departments were collated, and individual audit teams were contacted to verify the data accuracy. Audit teams failing to complete the full audit cycle with a re-audit were asked to explain the underlying reasons behind this.

Findings: Of the six trusts approached, two refused to participate, and one failed to initiate the project. A total of 39 audits were registered across three surgical directorates. Only 15 out of 39 audits completed at least one audit cycle, with 4 deemed of no value to re-audit. Only seven audits were completed to re-audit. Achieving a publication or a presentation was the most cited reason for not completing the audit loop.

Originality/value: This study demonstrates that the poor rates of audit completion rate found in other areas of clinical medicine pervade general surgery. Improved completion of an audit is essential and strategies to achieve this are urgently needed.
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http://dx.doi.org/10.1108/IJHCQA-06-2017-0109DOI Listing
October 2018

CD200 Expression Marks a Population of Quiescent Limbal Epithelial Stem Cells with Holoclone Forming Ability.

Stem Cells 2018 11 1;36(11):1723-1735. Epub 2018 Oct 1.

Institute of Genetic Medicine, Newcastle University, Newcastle, United Kingdom.

One of the main challenges in limbal stem cell (LSC) biology and transplantation is the lack of definitive cell surface markers which can be used to identify and enrich viable LSCs. In this study, expression of 361 cell surface proteins was assessed in ex vivo expanded limbal epithelial cells. One marker, CD200 was selected for further characterization based on expression in a small subset of limbal epithelial cells (2.25% ± 0.69%) and reduced expression through consecutive passaging and calcium induced differentiation. CD200 was localized to a small population of cells at the basal layer of the human and mouse limbal epithelium. CD200 cells were slow cycling and contained the majority of side population (SP) and all the holoclone forming progenitors. CD200 cells displayed higher expression of LSCs markers including PAX6, WNT7A, CDH3, CK14, CK15, and ABCB5 and lower expression of Ki67 when compared to CD200 . Downregulation of CD200 abrogated the ability of limbal epithelial cells to form holoclones, suggesting an important function for CD200 in the maintenance and/or self-renewal of LSCs. A second marker, CD109, which was expressed in 56.29% ± 13.96% of limbal epithelial cells, was also found to co-localize with ΔNp63 in both human and mouse cornea, albeit more abundantly than CD200. CD109 expression decreased slowly through calcium induced cell differentiation and CD109 cells were characterized by higher expression of Ki67, when compared to CD109 subpopulation. Together our data suggest that CD200 expression marks a quiescent population of LSCs with holoclone forming potential, while CD109 expression is associated with a proliferative progenitor phenotype. Stem Cells 2018;36:1723-1735.
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http://dx.doi.org/10.1002/stem.2903DOI Listing
November 2018

Clinical evaluation of Roche COBAS AmpliPrep/COBAS TaqMan CMV test using nonplasma samples.

J Med Virol 2018 10 12;90(10):1611-1619. Epub 2018 Jun 12.

Molecular Diagnostics Laboratory, Universtiy of Maryland Medical Center, Baltimore, Maryland.

Cytomegalovirus (CMV) infection is a leading cause of loss of hearing, vision, and mental retardation in congenitally infected children. It is also associated with complications of organ transplant and opportunistic HIV coinfection. The Roche COBAS AmpliPrep/COBAS TaqMan CMV test is an FDA-approved test that measures CMV DNA viral load in plasma for the diagnosis and management of patients at risk of CMV-associated diseases. Besides plasma, CMV is often found in bronchoalveolar lavage (BAL), cerebrospinal fluid (CSF), and urine. Thus, monitoring of CMV for critical care of patients in these nonplasma samples becomes necessary. The objective of this study was to conduct an analytic and clinical feasibility study of the Roche CMV test in BAL, CSF, and urine. The lower limit of detection, analytic measurement range, assay sensitivity, specificity, and precision were determined. Results of this study showed that the lower limit of detections were 50, 100, and 300 IU/mL for BAL, CSF, or urine, respectively. The analytic measurement ranges were from log 2.48 to log 5.48. The assay specificity was 94.4% for BAL and 100% for CSF and urine. The assay precision was all within the acceptable range. The performance of Roche test was further compared with 2 comparators including the RealTime CMV assay (Abbott Molecular) and a CMV Quantitative Polymerase Chain Reaction test (Vela Diagnostics). There was a general positive correlation between the Roche method and the Abbott or the Vela method. Overall, this study suggests that the Roche CMV test is suitable for the quantification of CMV viral load DNA in the described nonplasma samples.
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http://dx.doi.org/10.1002/jmv.25226DOI Listing
October 2018

Antibiotics versus no antibiotics in the treatment of acute uncomplicated diverticulitis - a systematic review and meta-analysis.

Colorectal Dis 2018 Jan 11. Epub 2018 Jan 11.

Royal Liverpool Hospital NHS Foundation Trust, Liverpool, UK.

Background: Acute uncomplicated diverticulitis (AUD) is common and antibiotics are the cornerstone of traditional conservative management. This approach lacks clear evidence base and studies have recently suggested that avoidance of antibiotics is a safe and efficacious way to manage AUD. The aim of this systematic review is to determine the safety and efficacy of treating AUD without antibiotics.

Methods: A systematic search of Embase, Cochrane library, MEDLINE, Science Citation Index Expanded, and ClinicalTrials. gov was performed. Studies comparing antibiotics versus no antibiotics in the treatment of AUD were included. Meta-analysis was performed using the random effects model with the primary outcome measure being diverticulitis-associated complications. Secondary outcomes were readmission rate, diverticulitis recurrence, mean hospital stay, requirement for surgery and requirement for percutaneous drainage.

Results: Eight studies were included involving 2469 patients; 1626 in the non-antibiotic group (NAb) and 843 in the antibiotic group (Ab). There was a higher complication rate in the Ab group however this was not significant (1.9% versus 2.6%) with a combined risk ratio (RR) of 0.63 (95% CI, 0.25 to 1.57, p=0.32). There was a shorter mean length of hospital stay in the Nab group (standard mean difference of -1.18 (95% CI, -2.34 to -0.03 p= 0.04). There was no significant difference in readmission, recurrence and surgical intervention rate or requirement for percutaneous drainage.

Conclusion: Treatment of AUD without antibiotics may be feasible with outcomes that are comparable to antibiotic treatment and with potential benefits for patients and the NHS. Large scale randomised multicentre studies are needed. This article is protected by copyright. All rights reserved.
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http://dx.doi.org/10.1111/codi.14013DOI Listing
January 2018

Dose Escalation Using Contact X-ray Brachytherapy After External Beam Radiotherapy as Nonsurgical Treatment Option for Rectal Cancer: Outcomes From a Single-Center Experience.

Int J Radiat Oncol Biol Phys 2018 03 20;100(3):565-573. Epub 2017 Oct 20.

Institute of Translational Medicine, University of Liverpool, Liverpool, United Kingdom; The Royal Liverpool and Broadgreen University Hospital National Health Service Trust, Liverpool, United Kingdom.

Purpose: To review the outcomes of rectal cancer patients treated with a nonsurgical approach using contact x-ray brachytherapy (CXB) when suspicious residual disease (≤3 cm) was present after external beam chemoradiation therapy/radiation therapy (EBCRT/EBRT).

Methods And Materials: Outcome data for rectal cancer patients referred to our institution from 2003 to 2012 were retrieved from an institutional database. These patients were referred after initial local multidisciplinary team discussion because they were not suitable for, or had refused, surgery. All selected patients received a CXB boost after EBCRT/EBRT. Most patients received a total of 90 Gy of CXB delivered in 3 fractions over 4 weeks.

Results: The median follow-up period was 2.5 years (range 1.2-8.3). Of 345 consecutive patients with rectal cancer referred to us, 83 with suspicious residual disease (≤3 cm) after EBCRT/EBRT were identified for a CXB boost. Their median age was 72 years (range 36-87), and 58 (69.9%) were men. The initial tumor stages were cT2 (n = 28) and cT3 (n = 55), and 54.2% were node positive. A clinical complete response (cCR) was achieved in 53 patients (63.8%) after the CXB boost that followed EBCRT/EBRT. Of these 53 patients, 7 (13.2%) developed a relapse after achieving a cCR, and the 6 patients (11.6%) with nonmetastatic regrowth underwent salvage surgery (100%). At the end of the study period, 69 of 83 patients (83.1%) were cancer free.

Conclusions: Our data suggest that a CXB boost for selected patients with suspicious residual disease (≤3 cm) after EBCRT/EBRT can be offered as an alternative to radical surgery. In our series, patients with a sustained cCR had a low rate of local regrowth, and those with nonmetastatic regrowth could be salvaged successfully. This approach could provide an alternative treatment option for elderly or comorbid patients who are not suitable for surgery and those with rectal cancer who wish to avoid surgery.
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http://dx.doi.org/10.1016/j.ijrobp.2017.10.022DOI Listing
March 2018

Dose escalation using contact X-ray brachytherapy (Papillon) for rectal cancer: does it improve the chance of organ preservation?

Br J Radiol 2017 Dec 10;90(1080):20170175. Epub 2017 Nov 10.

2 Institute of Translational Medicine , University of Liverpool , UK.

Objective: A watch and wait policy for patients with a clinical complete response (cCR) after external beam chemoradiotherapy (EBCRT) for rectal cancer is an attractive option. However, approximately one-third of tumours will regrow, which requires surgical salvage for cure. We assessed whether contact X-ray brachytherapy (CXB) can improve organ preservation by avoiding surgery for local regrowth.

Methods: From our institutional database, we identified 200 of 573 patients treated by CXB from 2003 to 2012. Median age was 74 years (range 32-94), and 134 (67%) patients were males. Histology was confirmed in all patients and was staged using CT scan, MRI or endorectal ultrasound. All patients received combined CXB and EBCRT, except 17 (8.5%) who had CXB alone.

Results: Initial cCR was achieved in 144/200 (72%) patients. 38/56 (68%) patients who had residual tumour received immediate salvage surgery. 16/144 (11%) patients developed local relapse after cCR, and 124/144 (86%) maintained cCR. At median follow up of 2.7 years, 161 (80.5%) patients were free of cancer. The main late toxicity was bleeding (28%). Organ preservation was achieved in 124/200 (62%) patients.

Conclusion: Our data suggest that CXB can reduce local regrowth to 11% compared with around 30% after EBCRT alone. Organ preservation of 62% achieved was higher than reported in most published watch and wait studies. Advances in knowledge: CXB is a promising treatment option to avoid salvage surgery for local regrowth, which can improve the chance of organ preservation in patients who are not suitable for or refuse surgery.
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http://dx.doi.org/10.1259/bjr.20170175DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6047662PMC
December 2017

Development and characterisation of a low-concentration sodium dodecyl sulphate decellularised porcine dermis.

J Tissue Eng 2017 Jan-Dec;8:2041731417724011. Epub 2017 Aug 3.

Institute of Medical and Biological Engineering, University of Leeds, Leeds, UK.

The aim of this study was to adapt a proprietary decellularisation process for human dermis for use with porcine skin. Porcine skin was subject to: sodium chloride (1 M) to detach the epidermis, trypsin paste to remove hair follicles, peracetic acid (0.1% v/v) disinfection, washed in hypotonic buffer and 0.1% (w/v) sodium dodecyl sulphate in the presence of proteinase inhibitors followed by nuclease treatment. Cellular porcine skin, decellularised porcine and human dermis were compared using histology, immunohistochemistry, GSL-1 lectin (alpha-gal epitope) staining, biochemical assays, uniaxial tensile and in vitro cytotoxicity tests. There was no microscopic evidence of cells in decellularised porcine dermis. DNA content was reduced by 98.2% compared to cellular porcine skin. There were no significant differences in the biomechanical parameters studied or evidence of cytotoxicity. The decellularised porcine dermis retained residual alpha-gal epitope. Basement membrane collagen IV immunostaining was lost following decellularisation; however, laminin staining was retained.
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http://dx.doi.org/10.1177/2041731417724011DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5546651PMC
August 2017

Human decellularized bone scaffolds from aged donors show improved osteoinductive capacity compared to young donor bone.

PLoS One 2017 15;12(5):e0177416. Epub 2017 May 15.

Divsion of Cell Matrix Biology and Regenerative Medicine, School of Biological Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester, United Kingdom.

To improve the safe use of allograft bone, decellularization techniques may be utilized to produce acellular scaffolds. Such scaffolds should retain their innate biological and biomechanical capacity and support mesenchymal stem cell (MSC) osteogenic differentiation. However, as allograft bone is derived from a wide age-range, this study aimed to determine whether donor age impacts on the ability an osteoinductive, acellular scaffold produced from human bone to promote the osteogenic differentiation of bone marrow MSCs (BM-MSC). BM-MSCs from young and old donors were seeded on acellular bone cubes from young and old donors undergoing osteoarthritis related hip surgery. All combinations resulted in increased osteogenic gene expression, and alkaline phosphatase (ALP) enzyme activity, however BM-MSCs cultured on old donor bone displayed the largest increases. BM-MSCs cultured in old donor bone conditioned media also displayed higher osteogenic gene expression and ALP activity than those exposed to young donor bone conditioned media. ELISA and Luminex analysis of conditioned media demonstrated similar levels of bioactive factors between age groups; however, IGF binding protein 1 (IGFBP1) concentration was significantly higher in young donor samples. Additionally, structural analysis of old donor bone indicated an increased porosity compared to young donor bone. These results demonstrate the ability of a decellularized scaffold produced from young and old donors to support osteogenic differentiation of cells from young and old donors. Significantly, the older donor bone produced greater osteogenic differentiation which may be related to reduced IGFBP1 bioavailability and increased porosity, potentially explaining the excellent clinical results seen with the use of allograft from aged donors.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0177416PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5432108PMC
September 2017

Short-term evolution of Shiga toxin-producing O157:H7 between two food-borne outbreaks.

Microb Genom 2016 09 8;2(9):e000084. Epub 2016 Sep 8.

2​Division of Infection and Immunity, The Roslin Institute and Royal (Dick) School of Veterinary Studies, University of Edinburgh, EH25 9RG Roslin, UK.

Shiga toxin-producing (STEC) O157:H7 is a public health threat and outbreaks occur worldwide. Here, we investigate genomic differences between related STEC O157:H7 that caused two outbreaks, eight weeks apart, at the same restaurant. Short-read genome sequencing divided the outbreak strains into two sub-clusters separated by only three single-nucleotide polymorphisms in the core genome while traditional typing identified them as separate phage types, PT8 and PT54. Isolates did not cluster with local strains but with those associated with foreign travel to the Middle East/North Africa. Combined long-read sequencing approaches and optical mapping revealed that the two outbreak strains had undergone significant microevolution in the accessory genome with prophage gain, loss and recombination. In addition, the PT54 sub-type had acquired a 240 kbp multi-drug resistance (MDR) IncHI2 plasmid responsible for the phage type switch. A PT54 isolate had a general fitness advantage over a PT8 isolate in rich medium, including an increased capacity to use specific amino acids and dipeptides as a nitrogen source. The second outbreak was considerably larger and there were multiple secondary cases indicative of effective human-to-human transmission. We speculate that MDR plasmid acquisition and prophage changes have adapted the PT54 strain for human infection and transmission. Our study shows the added insights provided by combining whole-genome sequencing approaches for outbreak investigations.
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http://dx.doi.org/10.1099/mgen.0.000084DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5320650PMC
September 2016

Characterisation of a divergent progenitor cell sub-populations in human osteoarthritic cartilage: the role of telomere erosion and replicative senescence.

Sci Rep 2017 02 2;7:41421. Epub 2017 Feb 2.

Centre for NanoHealth, Swansea University Medical School, Swansea, UK.

In recent years it has become increasingly clear that articular cartilage harbours a viable pool of progenitor cells and interest has focussed on their role during development and disease. Analysis of progenitor numbers using fluorescence-activated sorting techniques has resulted in wide-ranging estimates, which may be the result of context-dependent expression of cell surface markers. We have used a colony-forming assay to reliably determine chondroprogenitor numbers in normal and osteoarthritic cartilage where we observed a 2-fold increase in diseased tissue (P  < 0.0001). Intriguingly, cell kinetic analysis of clonal isolates derived from single and multiple donors of osteoarthritic cartilage revealed the presence of a divergent progenitor subpopulation characterised by an early senescent phenotype. Divergent sub-populations displayed increased senescence-associated β-galactosidase activity, lower average telomere lengths but retained the capacity to undergo multi-lineage differentiation. Osteoarthritis is an age-related disease and cellular senescence is predicted to be a significant component of the pathological process. This study shows that although early senescence is an inherent property of a subset of activated progenitors, there is also a pool of progenitors with extended viability and regenerative potential residing within osteoarthritic cartilage.
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http://dx.doi.org/10.1038/srep41421DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5288717PMC
February 2017

Development of decellularized conjunctiva as a substrate for the ex vivo expansion of conjunctival epithelium.

J Tissue Eng Regen Med 2018 02 12;12(2):e973-e982. Epub 2017 Jun 12.

National Health Service Blood and Transplant Tissue Services, Speke, Liverpool, UK.

This study was performed to develop a method to decellularize human conjunctiva and to characterize the tissue in terms of its deoxyribose nucleic acid (DNA) content, tensile strength, collagen denaturation, basement membrane, extracellular matrix components and its potential to support conjunctival epithelial growth. Human conjunctival tissues were subjected to a decellularization process involving hypotonic detergent and nuclease buffers. Variations in sodium dodecyl sulfate concentration (0.05-0.5%, w/v) were tested to determine the appropriate concentration of detergent buffer. DNA quantification, collagen denaturation, cytotoxicity and tensile strength were investigated. Human conjunctival cell growth by explant culture on the decellularized tissue substrate was assessed after 28 days in culture. Samples were fixed and paraffin embedded for immunohistochemistry including conjunctival epithelial cell markers and extracellular matrix proteins. Conjunctival tissue from 20 eyes of 10 donors (age range 65-92 years) was used. Decellularization of human conjunctiva was achieved to 99% or greater DNA removal (p < 0.001) with absence of nuclear staining. This was reproducible at the lowest concentration of sodium dodecyl sulfate (0.05% w/v). No collagen denaturation (p = 0.74) and no difference in tensile strength parameters was demonstrated following decellularization. No significant difference was noted in the immunolocalization of collagen IV, laminin and fibronectin, or in the appearance of periodic acid-Schiff-stained basement membranes following decellularization. The decellularized tissue did not exhibit any cytotoxicity and explant culture resulted in the growth of stratified conjunctival epithelium. Allogeneic decellularized human conjunctiva can be successfully decellularized using the described protocol. It represents a novel substrate to support the expansion of conjunctival epithelium for ocular surface cellular replacement therapies. Copyright © 2017 John Wiley & Sons, Ltd.
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http://dx.doi.org/10.1002/term.2419DOI Listing
February 2018

Pregnancy after abdominal wall mesh repair in desmoid fibromatosis.

J Obstet Gynaecol 2017 Apr 22;37(3):379-380. Epub 2016 Dec 22.

b Royal Liverpool and Broadgreen University Hospitals NHS Trust , Liverpool , UK.

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http://dx.doi.org/10.1080/01443615.2016.1250729DOI Listing
April 2017

Decellularization of human donor aortic and pulmonary valved conduits using low concentration sodium dodecyl sulfate.

J Tissue Eng Regen Med 2018 02 12;12(2):e841-e853. Epub 2017 May 12.

School of Biomedical Sciences, Faculty of Biological Sciences, University of Leeds, Leeds, UK.

The clinical use of decellularized cardiac valve allografts is increasing. Long-term data will be required to determine whether they outperform conventional cryopreserved allografts. Valves decellularized using different processes may show varied long-term outcomes. It is therefore important to understand the effects of specific decellularization technologies on the characteristics of donor heart valves. Human cryopreserved aortic and pulmonary valved conduits were decellularized using hypotonic buffer, 0.1% (w/v) sodium dodecyl sulfate and nuclease digestion. The decellularized tissues were compared to cellular cryopreserved valve tissues using histology, immunohistochemistry, quantitation of total deoxyribose nucleic acid, collagen and glycosaminoglycan content, in vitro cytotoxicity assays, uniaxial tensile testing and subcutaneous implantation in mice. The decellularized tissues showed no histological evidence of cells or cell remnants and >97% deoxyribose nucleic acid removal in all regions (arterial wall, muscle, leaflet and junction). The decellularized tissues retained collagen IV and von Willebrand factor staining with some loss of fibronectin, laminin and chondroitin sulfate staining. There was an absence of major histocompatibility complex Class I staining in decellularized pulmonary valve tissues, with only residual staining in isolated areas of decellularized aortic valve tissues. The collagen content of the tissues was not decreased following decellularization however the glycosaminoglycan content was reduced. Only moderate changes in the maximum load to failure of the tissues were recorded postdecellularization. The decellularized tissues were noncytotoxic in vitro, and were biocompatible in vivo in a mouse subcutaneous implant model. The decellularization process will now be translated into a good manufacturing practices-compatible process for donor cryopreserved valves with a view to future clinical use. Copyright © 2016 The Authors Tissue Engineering and Regenerative Medicine published by John Wiley & Sons, Ltd.
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http://dx.doi.org/10.1002/term.2391DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5836965PMC
February 2018

An important role for adenine, cholera toxin, hydrocortisone and triiodothyronine in the proliferation, self-renewal and differentiation of limbal stem cells in vitro.

Exp Eye Res 2016 11 28;152:113-122. Epub 2016 Sep 28.

Institute of Genetic Medicine, Newcastle University, UK. Electronic address:

The cornea is a self-renewing tissue located at the front of the eye. Its transparency is essential for allowing light to focus onto the retina for visual perception. The continuous renewal of corneal epithelium is supported by limbal stem cells (LSCs) which are located in the border region between conjunctiva and cornea known as the limbus. Ex vivo expansion of LSCs has been successfully applied in the last two decades to treat patients with limbal stem cell deficiency (LSCD). Various methods have been used for their expansion, yet the most widely used culture media contains a number of ingredients derived from animal sources which may compromise the safety profile of human LSC transplantation. In this study we sought to understand the role of these components namely adenine, cholera toxin, hydrocortisone and triiodothyronine with the aim of re-defining a safe and GMP compatible minimal media for the ex vivo expansion of LSCs on human amniotic membrane. Our data suggest that all four components play a critical role in maintaining LSC proliferation and promoting LSC self-renewal. However removal of adenine and triiodothyronine had a more profound impact and led to LSC differentiation and loss of viability respectively, suggesting their essential role for ex vivo expansion of LSCs. Replacement of each of the components with GMP-grade reagents resulted in equal growth to non-GMP grade media, however an enhanced differentiation of LSCs was observed, suggesting that additional combinations of GMP grade reagents need to be tested to achieve similar or better level of LSC maintenance in the same manner as the traditional LSC media.
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http://dx.doi.org/10.1016/j.exer.2016.09.008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5105828PMC
November 2016

Polyurethane scaffolds seeded with CD34(+) cells maintain early stem cells whilst also facilitating prolonged egress of haematopoietic progenitors.

Sci Rep 2016 08 30;6:32149. Epub 2016 Aug 30.

School of Biochemistry, Biomedical Sciences Building, University of Bristol, Bristol, BS8 1TD, United Kingdom.

We describe a 3D erythroid culture system that utilises a porous polyurethane (PU) scaffold to mimic the compartmentalisation found in the bone marrow. PU scaffolds seeded with peripheral blood CD34(+) cells exhibit a remarkable reproducibility of egress, with an increased output when directly compared to human bone scaffolds over 28 days. Immunofluorescence demonstrated the persistence of CD34(+) cells within the scaffolds for the entirety of the culture. To characterise scaffold outputs, we designed a flow cytometry panel that utilises surface marker expression observed in standard 2D erythroid and megakaryocyte cultures. This showed that the egress population is comprised of haematopoietic progenitor cells (CD36(+)GPA(-/low)). Control cultures conducted in parallel but in the absence of a scaffold were also generally maintained for the longevity of the culture albeit with a higher level of cell death. The harvested scaffold egress can also be expanded and differentiated to the reticulocyte stage. In summary, PU scaffolds can behave as a subtractive compartmentalised culture system retaining and allowing maintenance of the seeded "CD34(+) cell" population despite this population decreasing in amount as the culture progresses, whilst also facilitating egress of increasingly differentiated cells.
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http://dx.doi.org/10.1038/srep32149DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5004174PMC
August 2016

Yield and Viability of Human Limbal Stem Cells From Fresh and Stored Tissue.

Invest Ophthalmol Vis Sci 2016 Jul;57(8):3708-13

Department of Eye and Vision Science, University of Liverpool, Liverpool, United Kingdom 2St. Paul's Eye Unit, Royal Liverpool University Hospital, Liverpool, United Kingdom.

Purpose: We compared cell number, putative stem cell markers, and clonogenic ability in fresh uncultured human limbal epithelial cells to that obtained from stored organ-cultured tissue.

Methods: Cell suspensions were formed from fresh and organ culture-stored human limbal epithelium. Expression of putative stem cell markers ΔNp63 and TrkA was performed using immunofluorescent staining before culture. Colony-forming efficiency (CFE) assays were performed at first passage. The effects of tissue storage, age, and postmortem/culture times were analyzed in a general linear model.

Results: Limbal tissue from 94 donors (34 fresh and 60 stored) was compared. Three times more cells were obtained per eye from fresh (35.34 × 104; SD, 17.39) than stored (11.24 × 104; SD, 11.57; P < 0.01) tissue. A higher proportion of cells from fresh tissue were viable (91.9%; SD, 5.7 vs. 85%; SD, 10.8) P < 0.01. Higher total cell expression of ΔNp63 (20.19 × 104; SD, 15.5 vs. 3.28 104; SD, 4.33) and TrkA (59.24 × 104; SD, 13.21 vs. 7.65 × 104; SD, 1.05) was observed in fresh than stored tissue (P < 0.01). Colony-forming efficiency was higher for fresh (1.42; SD, 0.12) than stored (0.43; SD, 0.15; P < 0.01) cells. For stored tissue only, there was a significant inverse relationship between donor age and total number of cells isolated (R2 = 0.27, P < 0.001).

Conclusions: Storage of corneoscleral discs in organ culture medium leads to significant reduction in limbal epithelial cell number, expression of ΔNp63 and TrkA, and viability compared to fresh tissue. There is a smaller basal stem cell population in stored compared to fresh tissue.
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http://dx.doi.org/10.1167/iovs.16-19354DOI Listing
July 2016

Does MRI Restaging of Rectal Cancer After Chemoradiotherapy Actually Permit a Change in Surgical Management?

Ann Surg 2017 12;266(6):e104-e105

Department of Surgery, The Royal Liverpool and Broadgreen University Hospitals NHS Trust, Prescot Street, Liverpool, UK Department of Radiology, The Royal Liverpool and Broadgreen University Hospitals NHS Trust, Prescot Street, Liverpool, UK.

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http://dx.doi.org/10.1097/SLA.0000000000001789DOI Listing
December 2017

Bi-linear mechanical property determination of acellular human patellar tendon grafts for use in anterior cruciate ligament replacement.

J Biomech 2016 06 30;49(9):1607-1612. Epub 2016 Mar 30.

(IMBE) Institute of Medical and Biological Engineering, School of Mechanical Engineering, University of Leeds, Leeds, UK.

Anterior cruciate ligament rupture is rising in its prevalence amongst the young and those with physically active lifestyles. Acellular human patellar tendon (PT) grafts offer a promising restoration solution, returning knee joint stability and overcoming some of the current disadvantages of autologous or allogeneic grafts. However, it is necessary to ensure that the decellularisation bio-processes involved do not cause structural changes in the microstructure of the tendon tissue that may adversely affect the mechanical properties, particularly with respect to the physiological range of loading. Sixteen cadaveric human PT grafts were sourced and processed from eight donors, with full ethical approval and consent for use in research. Eight specimens were allocated for decellularisation, while the remaining eight contralateral specimens were used as native controls. Testing consisted of 12 preconditioning cycles followed by uniaxial extension until failure occurred. Stress-strain data was then fitted to a bi-linear model using least squares regression by a custom-written Matlab script. The elastic moduli for the toe region and linear region of each specimen were determined, in addition to the transition point co-ordinates and strain energy density for increasing strain. No significant differences were found between groups for all of the parameters investigated. Hence, the shape and magnitude of the stress-strain profile was found to be the same for both groups throughout loading. The results of this study indicated that decellularisation appeared to have no effect on the material properties of human PT grafts under quasistatic conditions. Therefore, acellular human PT grafts can offer a viable additional solution for ACL replacement compared to current autologous and allogeneic treatment options.
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http://dx.doi.org/10.1016/j.jbiomech.2016.03.041DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5756535PMC
June 2016

Human limbal mesenchymal stem cells express ABCB5 and can grow on amniotic membrane.

Regen Med 2016 Apr 11;11(3):273-86. Epub 2016 Mar 11.

Institute of Genetic Medicine, Faculty of Medical Sciences, Newcastle University, Newcastle Upon-Tyne, NE1 3BZ, UK.

Aim: To isolate and characterize limbal mesenchymal stem cells (LMSCs) from human corneoscleral rings.

Materials & Methods: Cells were isolated from corneoscleral rings and cultured in a mesenchymal stem cell (MSC)-selective media and examined for differentiation, phenotyping and characterization.

Results: LMSCs were capable of trilineage differentiation, adhered to tissue culture plastic, expressed HLA class I and cell surface antigens associated with human MSC while having no/low expression of HLA class II and negative hematopoietic lineage markers. They were capable for CXCL12-mediated cellular migration. LMSCs adhered, proliferated on amniotic membrane and expressed the common putative limbal stem cell markers.

Conclusion: Limbal-derived MSC exhibited plasticity, could maintain limbal markers expression and demonstrated viable growth on amniotic membrane.
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http://dx.doi.org/10.2217/rme-2016-0009DOI Listing
April 2016

Watch-and-wait approach versus surgical resection after chemoradiotherapy for patients with rectal cancer (the OnCoRe project): a propensity-score matched cohort analysis.

Lancet Oncol 2016 Feb 17;17(2):174-183. Epub 2015 Dec 17.

Department of Colorectal Surgery, The Christie NHS Foundation Trust, Manchester, UK.

Background: Induction of a clinical complete response with chemoradiotherapy, followed by observation via a watch-and-wait approach, has emerged as a management option for patients with rectal cancer. We aimed to address the shortage of evidence regarding the safety of the watch-and-wait approach by comparing oncological outcomes between patients managed by watch and wait who achieved a clinical complete response and those who had surgical resection (standard care).

Methods: Oncological Outcomes after Clinical Complete Response in Patients with Rectal Cancer (OnCoRe) was a propensity-score matched cohort analysis study, that included patients of all ages diagnosed with rectal adenocarcinoma without distant metastases who had received preoperative chemoradiotherapy (45 Gy in 25 daily fractions with concurrent fluoropyrimidine-based chemotherapy) at a tertiary cancer centre in Manchester, UK, between Jan 14, 2011, and April 15, 2013. Patients who had a clinical complete response were offered management with the watch-and-wait approach, and patients who did not have a complete clinical response were offered surgical resection if eligible. We also included patients with a clinical complete response managed by watch and wait between March 10, 2005, and Jan 21, 2015, across three neighbouring UK regional cancer centres, whose details were obtained through a registry. For comparative analyses, we derived one-to-one paired cohorts of watch and wait versus surgical resection using propensity-score matching (including T stage, age, and performance status). The primary endpoint was non-regrowth disease-free survival from the date that chemoradiotherapy was started, and secondary endpoints were overall survival, and colostomy-free survival. We used a conservative p value of less than 0·01 to indicate statistical significance in the comparative analyses.

Findings: 259 patients were included in our Manchester tertiary cancer centre cohort, 228 of whom underwent surgical resection at referring hospitals and 31 of whom had a clinical complete response, managed by watch and wait. A further 98 patients were added to the watch-and-wait group via the registry. Of the 129 patients managed by watch and wait (median follow-up 33 months [IQR 19-43]), 44 (34%) had local regrowths (3-year actuarial rate 38% [95% CI 30-48]); 36 (88%) of 41 patients with non-metastatic local regrowths were salvaged. In the matched analyses (109 patients in each treatment group), no differences in 3-year non-regrowth disease-free survival were noted between watch and wait and surgical resection (88% [95% CI 75-94] with watch and wait vs 78% [63-87] with surgical resection; time-varying p=0·043). Similarly, no difference in 3-year overall survival was noted (96% [88-98] vs 87% [77-93]; time-varying p=0·024). By contrast, patients managed by watch and wait had significantly better 3-year colostomy-free survival than did those who had surgical resection (74% [95% CI 64-82] vs 47% [37-57]; hazard ratio 0·445 [95% CI 0·31-0·63; p<0·0001), with a 26% (95% CI 13-39) absolute difference in patients who avoided permanent colostomy at 3 years between treatment groups.

Interpretation: A substantial proportion of patients with rectal cancer managed by watch and wait avoided major surgery and averted permanent colostomy without loss of oncological safety at 3 years. These findings should inform decision making at the outset of chemoradiotherapy.

Funding: Bowel Disease Research Foundation.
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http://dx.doi.org/10.1016/S1470-2045(15)00467-2DOI Listing
February 2016

Evaluation of Copper and Hydrogen Peroxide Treatments on the Biology, Biomechanics, and Cytotoxicity of Decellularized Dermal Allografts.

Tissue Eng Part C Methods 2016 Mar 29;22(3):290-300. Epub 2016 Jan 29.

NHS Blood and Transplant, Tissue Services R&D , Liverpool, United Kingdom .

Decellularized tissue allografts are paving the way as an alternative to cellular tissue transplantation. Effective sterilization or decontamination of tissue allografts is paramount for the safety of the allograft; however, some of the current sterilization procedures have a detrimental effect on the tissue scaffold. The bactericidal and virucidal activity of copper (II) ions and hydrogen peroxide (H2O2) have been widely reported, however, their effect on the biology, biochemistry, and biocompatibility of decellularized tissue have yet to be elucidated. In this study, decellularized human dermis (dCELL human dermis) was treated with copper (II) chloride (CuCl2) and H2O2; both singly and in combination, and parameters, including concentration, pH, and synergy between CuCl2 and H2O2, were evaluated to identify conditions where any detrimental effects on the tissue scaffold were observed. Skin from 13 human donors was retrieved with appropriate consent and processed into dCELL human dermis. The dCELL human dermis was then treated for 3 h with 0.1 mg/L-1 g/L (w/v) CuCl2 and 0.01-7.5% (v/v) H2O2 and combinations of both of these in the same concentration range. dCELL human dermis treated with solutions of 0.1 mg/L-1 g/L CuCl2 or 0.01-7.5% H2O2 caused no detrimental effects on gross histology, collagen denaturation, collagen orientation, and biomechanical properties of the tissue or cytotoxicity. The highest combined concentration of CuCl2 and H2O2 demonstrated an increase in ultimate tensile strength, loss of collagen type IV immunostaining at the dermal-epidermal junction, and in vitro cytotoxicity. Combinations within the range of up to 10 mg/L CuCl2 with up to 0.5% H2O2 had no effect. The data identify the concentrations of CuCl2 and H2O2 solutions that have no effect on the biological, biomechanical, and biochemical properties of dCELL human dermis, while retaining biocompatibility. These treatments may be suitable for use as sterilization/decontamination agents on human decellularized tissues.
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http://dx.doi.org/10.1089/ten.TEC.2015.0271DOI Listing
March 2016

Adjuvant chemotherapy for rectal cancer: Is it needed?

World J Clin Oncol 2015 Dec;6(6):225-36

Kristijonas Milinis, Faculty of Medicine, University of Liverpool, Liverpool L69 3GE, United Kingdom.

Adjuvant chemotherapy has become a standard treatment of advanced rectal cancer in the West. The benefits of adjuvant chemotherapy after surgery alone have been well established. However, controversy surrounds the use adjuvant chemotherapy in patients who received preoperative chemoradiotherapy, despite it being recommended by a number of international guidelines. Results of recent multicentre randomised control trials showed no benefit of adjuvant chemotherapy in terms of survival and rates of distant metastases. However, concerns exist regarding the quality of the studies including inadequate staging modalities, out-dated chemotherapeutic regimens and surgical approaches and small sample sizes. It has become evident that not all the patients respond to adjuvant chemotherapy and more personalised approach should be employed when considering the benefits of adjuvant chemotherapy. The present review discusses the strengths and weaknesses of the current evidence-base and suggests improvements for future studies.
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http://dx.doi.org/10.5306/wjco.v6.i6.225DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4675908PMC
December 2015

Short article: Successful faecal coliform sensitivity-based oral ertapenem therapy for chronic antibiotic-refractory pouchitis: a case series.

Eur J Gastroenterol Hepatol 2016 Mar;28(3):277-80

Departments of aGastroenterology bColorectal Surgery cPathology dMedical Microbiology, Royal Liverpool University Hospital, Liverpool, UK.

Pouchitis is a common complication of restorative proctocolectomy for ulcerative colitis, and a proportion of patients develop a refractory course. The treatment of chronic antibiotic-refractory pouchitis (CARP) is challenging, and treatment failure is often a cause of pouch excision. We report on a series of three patients with CARP who were treated with oral ertapenem following faecal coliform sensitivity testing. There was an improvement in the pouchitis disease activity index in all three patients [pretreatment pouch disease activity index, median 13 (range: 10-14); post-treatment pouch disease activity index, median 1 (range: 1-3)]. Identification of faecal coliform sensitivity and treatment with oral ertapenem might be helpful in patients with CARP.
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http://dx.doi.org/10.1097/MEG.0000000000000549DOI Listing
March 2016