Identification of Intervention Opportunities through Assessment of the Appropriateness of Antibiotic Prescribing in Surgical Patients in a UK Hospital Using a National Audit Tool: A Single Centre Retrospective Audit.

Identifying opportunities to safely reduce antibiotic prescribing is necessary for prescribers and antibiotic stewardship teams to minimise unwarranted antibiotic use. We aimed to quantify excess antibiotic use in General Surgery. We retrospectively audited the antibiotic prescribing for patients discharged from the General Surgery specialty in an acute hospital in the south-west of England for one month using an audit tool developed by Public Health England. The appropriateness of prescribing was determined for each patient at three antibiotic decision time-points: at initiation, the pre-72-hour antibiotic review, and treatment duration. Two infection specialists and a general surgeon reviewed each patient. Indication and excess days of therapy (DOTs) were calculated at each decision time-point and expressed as a proportion of total DOTs. Eighty-six patients were prescribed 1162 DOTs; 192 (16.5%) excess DOTs were prescribed in 38 patients (44%), with zero excess days identified in the remaining 48 patients (56%). Seventy-five of 192 (39%) excess DOTs occurred at initiation; 55/192 (29%) after the pre-72-hour antibiotic review; and 62/192 (32%) due to protracted antibiotic courses. There was concordance between the general surgeon and infection specialist for most apportioned excess DOTs. However, the surgeon apportioned fewer excess DOTs 160/1162 (13.8%). Overall IV antibiotics accounted for 53.4% of total DOTs. Seventy-two of 86 (83.7%) patients received 620 intravenous DOTs; of these, 79 (12.7%) IV DOTS were unnecessary. We have identified excess antibiotic prescribing in General surgery with comparable excess DOTs at all three time-points.

Serial Clustering of Late-Onset Group B Streptococcal Infections in the Neonatal Unit: A Genomic Re-evaluation of Causality.

Background: Invasive Group B streptococcus (GBS) is a major cause of serious neonatal infection. Current strategies to reduce early-onset GBS disease have no impact on late-onset disease (LOD). Although GBS LOD is viewed as a sporadic event in the community, LOD arising within the neonatal intensive care unit (ICU) raises questions about mode of acquisition. Methods: Following a cluster of 4 GBS LOD cases, enhanced surveillance for all GBS LOD was undertaken over 2 years in the neonatal ICU supported by neonatal rectal screening. GBS isolates were serotyped and genome-sequenced. Results: Twelve late -onset invasive GBS episodes were identified (incidence 0.6/1000 live births). Genomic analysis revealed that 11/12 GBS isolates (92%) were linked to at least one other LOD isolate. Isolates from the first cluster were serotype V, resistant to macrolides and lincosamides, and sequencing confirmed isolates were indistinguishable, or distinguishable by only one SNP difference, from each other. Rectal carriage was rare. Prospective surveillance identified three further clusters of LOD due to serotypes Ia (3 cases), Ib (2 cases), and III (2 cases), that would not have been identified without surveillance and genome sequencing, leading to a re-evaluation of interventions required to prevent GBS LOD. Conclusion: Acquisition routes for LOD GBS in the neonatal ICU are poorly understood; cases may not necessarily be sporadic. Within this neonatal ICU, our data suggest that a single case of LOD GBS sepsis should be considered a potential nosocomial transmission event warranting prompt investigation, heightened infection prevention vigilance and action where required.

Emergence and clonal spread of colistin resistance due to multiple mutational mechanisms in carbapenemase-producing Klebsiella pneumoniae in London.

Carbapenemase-producing Enterobacteriaceae (CPE) are emerging worldwide, limiting therapeutic options. Mutational and plasmid-mediated mechanisms of colistin resistance have both been reported. The emergence and clonal spread of colistin resistance was analysed in 40 epidemiologically-related NDM-1 carbapenemase producing Klebsiella pneumoniae isolates identified during an outbreak in a group of London hospitals. Isolates from July 2014 to October 2015 were tested for colistin susceptibility using agar dilution, and characterised by whole genome sequencing (WGS). Colistin resistance was detected in 25/38 (65.8%) cases for which colistin susceptibility was tested. WGS found that three potential mechanisms of colistin resistance had emerged separately, two due to different mutations in mgrB, and one due to a mutation in phoQ, with onward transmission of two distinct colistin-resistant variants, resulting in two sub-clones associated with transmission at separate hospitals. A high rate of colistin resistance (66%) emerged over a 10 month period. WGS demonstrated that mutational colistin resistance emerged three times during the outbreak, with transmission of two colistin-resistant variants.

Cerebral Abscess Associated With Odontogenic Bacteremias, Hypoxemia, and Iron Loading in Immunocompetent Patients With Right-to-Left Shunting Through Pulmonary Arteriovenous Malformations.

Background: Cerebral abscess is a recognized complication of pulmonary arteriovenous malformations (PAVMs) that allow systemic venous blood to bypass the pulmonary capillary bed through anatomic right-to-left shunts. Broader implications and mechanisms remain poorly explored. Methods: Between June 2005 and December 2016, at a single institution, 445 consecutive adult patients with computed tomography-confirmed PAVMs (including 403 [90.5%] with hereditary hemorrhagic telangiectasia) were recruited to a prospective series. Multivariate logistic regression was performed and detailed periabscess histories were evaluated to identify potential associations with cerebral abscess. Rates were compared to an earlier nonoverlapping series. Results: Thirty-seven of the 445 (8.3%) patients experienced a cerebral abscess at a median age of 50 years (range, 19-76 years). The rate adjusted for ascertainment bias was 27 of 435 (6.2%). Twenty-nine of 37 (78.4%) patients with abscess had no PAVM diagnosis prior to their abscess, a rate unchanged from earlier UK series. Twenty-one of 37 (56.7%) suffered residual neurological deficits (most commonly memory/cognition impairment), hemiparesis, and visual defects. Isolation of periodontal microbes, and precipitating dental and other interventional events, emphasized potential sources of endovascular inoculations. In multivariate logistic regression, cerebral abscess was associated with low oxygen saturation (indicating greater right-to-left shunting); higher transferrin iron saturation index; intravenous iron use for anemia (adjusted odds ratio, 5.4 [95% confidence interval, 1.4-21.1]); male sex; and venous thromboemboli. There were no relationships with anatomic attributes of PAVMs, or red cell indices often increased due to secondary polycythemia. Conclusions: Greater appreciation of the risk of cerebral abscess in undiagnosed PAVMs is required. Lower oxygen saturation and intravenous iron may be modifiable risk factors.

Staphylococcus aureus adapts to oxidative stress by producing H2O2-resistant small-colony variants via the SOS response.

The development of chronic and recurrent Staphylococcus aureus infections is associated with the emergence of slow-growing mutants known as small-colony variants (SCVs), which are highly tolerant of antibiotics and can survive inside host cells. However, the host and bacterial factors which underpin SCV emergence during infection are poorly understood. Here, we demonstrate that exposure of S. aureus to sublethal concentrations of H2O2 leads to a specific, dose-dependent increase in the population frequency of gentamicin-resistant SCVs. Time course analyses revealed that H2O2 exposure caused bacteriostasis in wild-type cells during which time SCVs appeared spontaneously within the S. aureus population. This occurred via a mutagenic DNA repair pathway that included DNA double-strand break repair proteins RexAB, recombinase A, and polymerase V. In addition to triggering SCV emergence by increasing the mutation rate, H2O2 also selected for the SCV phenotype, leading to increased phenotypic stability and further enhancing the size of the SCV subpopulation by reducing the rate of SCV reversion to the wild type. Subsequent analyses revealed that SCVs were significantly more resistant to the toxic effects of H2O2 than wild-type bacteria. With the exception of heme auxotrophs, gentamicin-resistant SCVs displayed greater catalase activity than wild-type bacteria, which contributed to their resistance to H2O2. Taken together, these data reveal a mechanism by which S. aureus adapts to oxidative stress via the production of a subpopulation of H2O2-resistant SCVs with enhanced catalase production.

Harnessing the genome: development of a hierarchical typing scheme for meticillin-resistant Staphylococcus aureus.

A major barrier to using genome sequencing in medical microbiology is the ability to interpret the data. New schemes that provide information about the importance of sequence variation in both clinical and public health settings are required. Meticillin-resistant Staphylococcus aureus (MRSA) is an important nosocomial pathogen that is being observed with increasing frequency in community settings. Better tools are needed to improve our understanding of its transmissibility and micro-epidemiology in order to develop effective interventions. Using DNA microarray technology we identified a set of 20 binary targets whose presence or absence could be determined by PCR, producing a PCR binary typing scheme (PCR-BT). This was combined with multi-locus sequence type-based, sequence nucleotide polymorphism typing to form a hierarchical typing scheme. When applied to a set of epidemiologically unrelated isolates, a high degree of concordance was observed with PFGE (98.8 %). The scheme was able to detect the presence or absence of an outbreak strain in eight out of nine outbreak investigations, demonstrating epidemiological concordance. PCR-BT was better than PFGE at distinguishing between outbreak strains, particularly where epidemic MRSA-15 was involved. The method developed here is a rapid, digital typing scheme for S. aureus for use in both micro- and macro-epidemiological investigations that has the advantage of being suitable for use in routine diagnostic laboratories. The targets are defined and therefore the types can be defined by any platform capable of detecting the sequences used, including whole genome sequencing.

Emergence and global spread of epidemic healthcare-associated Clostridium difficile.

Epidemic C. difficile (027/BI/NAP1) has rapidly emerged in the past decade as the leading cause of antibiotic-associated diarrhea worldwide. However, the key events in evolutionary history leading to its emergence and the subsequent patterns of global spread remain unknown. Here, we define the global population structure of C. difficile 027/BI/NAP1 using whole-genome sequencing and phylogenetic analysis. We show that two distinct epidemic lineages, FQR1 and FQR2, not one as previously thought, emerged in North America within a relatively short period after acquiring the same fluoroquinolone resistance-conferring mutation and a highly related conjugative transposon. The two epidemic lineages showed distinct patterns of global spread, and the FQR2 lineage spread more widely, leading to healthcare-associated outbreaks in the UK, continental Europe and Australia. Our analysis identifies key genetic changes linked to the rapid transcontinental dissemination of epidemic C. difficile 027/BI/NAP1 and highlights the routes by which it spreads through the global healthcare system.

Clinical trials of GMP products in the gene therapy field.

Advances in gene therapy are increasingly leading to clinical assessment in many fields of medicine with diverse approaches. The basic science stems from approaches aimed at different functions such as correcting a missing/abnormal gene, altering the proportion or expression of normal genes to augment a physiological process or using this principle to destroy malignant or infected cells. As the technology advances, it is increasingly important to ensure that clinical trials answer the questions that need to be asked. In this chapter we review examples of published clinical trials, resources for accessing information about registered trials, the process of regulating trials, good clinical practice, and good manufacturing practice as well as summarising the approach taken by regulatory authorities in reviewing applications for the introduction of products for use in the clinic.

Helicobacter pylori stimulates dendritic cells to induce interleukin-17 expression from CD4+ T lymphocytes.

Helicobacter pylori is a human gastroduodenal pathogen that leads to active chronic inflammation characterized by T-cell responses biased toward a Th1 phenotype. It has been accepted that H. pylori infection induces a Th17 response. At mucosal sites, dendritic cells (DCs) have the capacity to induce effector T cells. Here, we evaluate the role of DCs in the H. pylori-induced interleukin-17 (IL-17) response. Immunohistochemistry and immunofluorescence were performed on human gastric mucosal biopsy samples and showed that myeloid DCs in H. pylori-infected patients colocalized with IL-23- and that IL-17-producing lymphocytes were present in H. pylori-infected antral biopsy samples. In parallel, human monocyte-derived DCs stimulated in vitro with live H. pylori cells produced significant levels of IL-23 in the absence of IL-12 release. The subsequent incubation of H. pylori-infected DCs with autologous CD4(+) T cells led to gamma interferon (IFN-gamma) and IL-17 expression. The inhibition of IL-1 and, to a lesser extent, IL-23 inhibited IL-17 production by T cells. Finally, isogenic H. pylori mutant strains not expressing major virulence factors were less effective in inducing IL-1 and IL-23 release by DCs and IL-17 release by T cells than parental strains. Altogether, we can conclude that DCs are potent inducers of IL-23/IL-17 expression following H. pylori stimulation. IL-1/IL-23 as well as H. pylori virulence factors seem to play an important role in mediating this response.

TLR9 polymorphisms determine murine lymphocyte responses to Helicobacter: results from a genome-wide scan.

Immune responses to microorganisms in the gastrointestinal tract must be carefully controlled to avoid disease. Helicobacter are Gram-negative bacteria which cause persistent infection and, in a minority of hosts, peptic ulceration or gastric cancer. Lymphocyte responses are important determinants of the outcome of infection. Therefore, it is important to identify the genetic determinants of lymphocyte responses to this mucosal pathogen. Using a (C57BL/6xBALB/c) F2 mouse model of Helicobacter infection, we mapped a region of linkage for lymphoproliferation to chromosome 9. Analysis of candidate genes in this region revealed variation of DNA sequence and gene expression in the TLR9 gene between C57BL/6 and BALB/c mouse strains. Reporter assays demonstrated higher levels of TLR9 transcriptional activity and increased NF-kappaB activation associated with the C57BL/6 TLR9 promoter and coding sequences. The importance of TLR9 in the control of lymphocyte responses was confirmed by demonstrating that lymphoproliferation and IFN-gamma secretion was diminished in the TLR9-/- mouse. Furthermore, neutrophil infiltration of the gastric epithelium is reduced in the absence of TLR9. Regulation of TLR9 expression and signalling therefore appears to play an important role in the control of lymphocyte responses to Helicobacter and potentially other luminal microorganisms.

Chronic exposure to Helicobacter pylori impairs dendritic cell function and inhibits Th1 development.

Helicobacter pylori causes chronic gastric infection that affects the majority of the world's population. Despite generating an inflammatory response, the immune system usually fails to clear the infection. Since dendritic cells (DCs) play a pivotal role in shaping the immune response, we investigated the effects of H. pylori on DC function. We have demonstrated that H. pylori increased the expression of activation markers on DCs while upregulating the inhibitory B7 family molecule, PD-L1. Functionally, H. pylori-treated DCs resulted in the production of interleukin-10 (IL-10) and IL-23 but not of alpha interferon (IFN-alpha). While very little or no IL-12 was produced to H. pylori alone, simultaneous ligation of CD40 on DCs induced IL-12 release. We also demonstrated that DCs treated with H. pylori-induced IFN-gamma production by allogeneic naive T cells. However, stimulation of DCs with H. pylori for an extended period of time impaired their ability to produce cytokines after CD40 ligation and limited their ability to promote IFN-gamma release, suggesting that the DCs had become exhausted by the prolonged stimulation. The effect of chronic infection with H. pylori on DC function was further investigated by focusing on DC development. Demonstrating that monocytes differentiated into DCs in the presence of H. pylori exhibited an exhausted phenotype with an impaired ability to produce IL-12 and a downregulation of CD1a. Our results raise the possibility that in chronic H. pylori infection DCs become exhausted after prolonged antigen exposure leading to suboptimal Th1 development. This effect may contribute to persistence of H. pylori infection.

A new approach to treatment of resistant gram-positive infections: potential impact of targeted IV to oral switch on length of stay.

BACKGROUND: Patients prescribed intravenous (IV) glycopeptides usually remain in hospital until completion of this treatment. Some of these patients could be discharged earlier if a switch to an oral antibiotic was made. This study was designed to identify the percentage of inpatients currently prescribed IV glycopeptides who could be discharged earlier if a switch to an oral agent was used, and to estimate the number of bed days that could be saved. We also aimed to identify the patient group(s) most likely to benefit, and to estimate the number of days of IV therapy that could be prevented in patients who remained in hospital. METHODS: Patients were included if they were prescribed an IV glycopeptide for 5 days or more. Predetermined IV to oral antibiotic switch criteria and discharge criteria were applied. A multiple logistic regression model was used to identify the characteristics of the patients most likely to be suitable for earlier discharge. RESULTS: Of 211 patients, 62 (29%) could have had a reduced length of stay if they were treated with a suitable oral antibiotic. This would have saved a total of 649 inpatient days (median 5 per patient; range 1-54). A further 31 patients (15%) could have switched to oral therapy as an inpatient thus avoiding IV line use. The patients most likely to be suitable for early discharge were those with skin and soft tissue infection, under the cardiology, cardiothoracic surgery, orthopaedics, general medical, plastic surgery and vascular specialities, with no high risk comorbidity and less than five other regularly prescribed drugs. CONCLUSION: The need for glycopeptide therapy has a significant impact on length of stay. Effective targeting of oral antimicrobials could reduce the need for IV access, allow outpatient treatment and thus reduce the length of stay in patients with infections caused by antibiotic resistant gram-positive bacteria.

Reducing dialysis associated bacteraemia, and recommendations for surveillance in the United Kingdom: prospective study.

PROBLEM: Bacteraemia in dialysis units accounts for major morbidity, mortality, and antibiotic usage. Risk is much greater when lines rather than fistulas are used for haemodialysis. Surveillance is critical for infection control, but no standardised surveillance scheme exists in the United Kingdom. DESIGN: Prospective study in a London dialysis unit of the implementation and applicability of a dialysis associated bacteraemia surveillance scheme developed in the United States and its effect on bacteraemia, antibiotic usage, and admission. SETTING: Hammersmith Hospital dialysis unit, London, where 112 outpatients receive dialysis three times weekly. Between June 2002 and December 2004, 3418 patient months of data were collected. KEY MEASURES FOR IMPROVEMENT: Successful adoption of the scheme and reductions in bacteraemia rates, antibiotic usage, and admission to hospital. Strategy for improvement Embedding the surveillance scheme in the unit's clinical activity. EFFECTS OF CHANGE: Raised awareness of bacteraemia prevention, prudent antibiotic prescribing, and the need for improved provision of vascular access. The scheme required two hours a month of consultant time. Significant downward trends were seen in bacteraemia rates and antibiotic usage: mean rate ratios from quarter to quarter 0.90 (95% confidence interval 0.85 to 0.94) and 0.91 (0.87 to 0.96), respectively. The rate of admission to hospital also showed a significant downward trend, with admissions directly connected to access related infection declining more rapidly: mean rate ratio of successive quarters 0.90 (0.84 to 0.96). The overall proportion of patients dialysed through catheters was significantly higher than in US outpatient centres (62.3% v 29.4%, P < 0.01). Study data were successfully used in a business case to improve access provision. LESSONS LEARNT: Dialysis specific surveillance of bacteraemia is critical to infection control in dialysis units and improving quality of care. Such a scheme could be adopted across the United Kingdom.

Deep-seated resistance in relapsed paratyphoid fever.

We describe a case of relapsed paratyphoid fever in which the isolate had reduced susceptibility to ciprofloxacin due to a rare mutation within the gyrA gene. 18fluorodeoxyglucose positron emission tomography scanning identified deep-seated infection including unsuspected aortitis and highlights the utility of novel imaging techniques to improve our understanding and treatment of this disease.

Helicobacter pylori has stimulatory effects on naive T cells.

BACKGROUND: Despite an apparently active host response, Helicobacter pylori infection can persist for life. Unexpectedly, T cells from apparently uninfected individuals respond to H. pylori antigen by proliferating. Also, the T-cell proliferative response appears to be less in infected compared with uninfected individuals. MATERIALS AND METHODS: We have investigated the T-cell response of isolated human peripheral blood, naive, and memory CD4+ T cells to H. pylori antigen in infected and uninfected subjects. RESULTS: In agreement with previous findings, the peripheral blood proliferative response was higher in uninfected compared with infected subjects. Interestingly, there was a response in CD4+ CD45RO+ (memory) and CD4+CD45RA+ (naive) subsets. The RO/RA ratio of the response to H. pylori antigen was 0.8-2.1 in both H. pylori-positive and H. pylori-negative subjects, which was similar to that of a known superantigen (2.5 and 2.2 in Helicobacter-positive and -negative subjects, respectively) whereas the RO/RA response ratio to a recall antigen (tetanus toxoid) was 9.8 and 18.7 in Helicobacter-positive and -negative subjects, respectively. Mononuclear cells isolated from cord blood also responded to H. pylori antigen, whereas there was no response to tetanus toxoid. The cord blood response and CD4+ CD45RA+ cell response to H. pylori antigen were inhibited predominantly by anti-HLA-DR and to some extent by anti-HLA-DQ antibodies. Investigation of the response to five different recombinant H. pylori antigens identified two that produced a response in naive T cells. CONCLUSIONS: These data suggest that H. pylori possesses molecules that cause higher than expected proliferation of naive T cells.

Qualitatively distinct patterns of cytokines are released by human dendritic cells in response to different pathogens.

Dendritic cells produce cytokines that regulate the class of the adaptive immune response. Microbial recognition is mediated, at least in part, by pattern recognition receptors such as Toll-like receptors, which influence dendritic cell maturation. In humans it is not yet clear how intact pathogens modulate the developing immune response. To address the effects of intact pathogens on the maturation and effector functions of human dendritic cells, we investigated their responses to a number of microbial pathogens. We studied a range of micro-organisms including Gram-negative bacteria (Escherichia coli and Salmonella enterica sv. typhimurium), Gram-positive cocci (Staphylococcus aureus) and atypical bacteria (Mycobacterium tuberculosis and Mycoplasma hominis) as well as the human protozoal parasite Trichomonas vaginalis. The micro-organisms were fixed in formaldehyde to prevent replication whilst preserving surface morphology. All the pathogens induced similar up-regulation of dendritic cell activation-associated cell surface markers but there was a profound difference in the patterns of cytokines produced by the stimulated dendritic cells. Some pathogens (E. coli, Salmonella enterica sv. typhimurium and S. aureus) induced interleukin-12 (IL-12), IL-10 and interferon-alpha whereas others (M. tuberculosis, Mycoplasma hominis and T. vaginalis) induced only IL-10. This differential effect was not altered by costimulation of the dendritic cells through CD40. These results support the notion that human dendritic cells are plastic in their response to microbial stimuli and that the nature of the pathogen dictates the response of the dendritic cell.

A case of Pasteurella multocida peritoneal dialysis-associated peritonitis and review of the literature.

OBJECTIVES: Two episodes of peritoneal dialysis-associated peritonitis, which occurred four months apart and were both due to Pasteurella multocida, were noted in a 73 year old woman. This report aims to describe the clinical history of these episodes and the microbiological investigations that were undertaken. The relevant literature will also be discussed. METHODS AND RESULTS: Basic microbiological tests identified the organism as Pasteurella multocida, and further work at a specialist laboratory classified it as Pasteurella multocida subsp. multocida. Pulsed field gel electrophoresis confirmed that the strains isolated from the two clinical episodes originated from the same clone. A literature search was performed, looking particularly for patients who experienced more than one episode of peritonitis caused by Pasteurella spp, whether due to recurrence or re-infection. CONCLUSIONS: It is likely that the infection resulted from a domestic cat, as there was evidence of a cat bite to the dialysis tubing in the period between the two episodes. Re-infection with two identical strains of pasteurella is more probable than relapse, for reasons discussed. Strict hygiene and avoiding contact between dialysis tubing and domestic animals must be emphasised to try to prevent pasteurella and other animal-associated infections in this already vulnerable population.