A study of colonisation of the human gastrointestinal tract by multidrug resistant Escherichia coli

Escherichia coli is a commensal organism recognised as a globally important pathogen as it can cause severe invasive disease, including meningitis and bacteraemia. E. coli is a known carrier of antimicrobial resistance (AMR) and is a major contributor to the global spread of AMR. Mobile genetic elements (MGEs), including plasmids and insertion sequence (IS) elements, are reported to play a crucial role in the spread of AMR and in the creation of multidrug resistant (MDR) organisms. The occurrence of MDR organisms, including MDR E. coli, is a cause of significant concern as it restricts treatment options, and increases patient morbidity and mortality. International travel and hospital admissions often result in increased colonisation with MDR organisms, and both are known to contribute to the global spread of AMR. Colonisation with drug resistant organisms is a potential route of AMR spread which has not been previously characterised during a traveller stay and there is limited information on colonisation in a hospital inpatient stay. Historical studies of clinical isolates have previously been used to demonstrate the impact of the discovery and increased use of antimicrobials on the rise of AMR, particularly in the last few decades. Previous historical studies have focussed on adult populations which are more likely to have had exposure to a wide range of antimicrobials over longer periods of time. This thesis is a study of E. coli in three different settings. It looks at MDR E. coli colonising healthy travellers to Laos and UK intensive care unit patients, and E. coli causing infection in Dutch neonates. The overall aim of this thesis was to understand the dynamics of colonisation with MDR organisms and the dynamics of MDR infection in the absence of antibiotic selection. Colonisation and infection with MDR E. coli both in the absence of antibiotic selection can improve understanding of AMR in general with no bias to a resistance driven by a specific antibiotic. Strain level sampling methods in combination with short and long read whole genome sequencing (WGS) were used to explore E. coli from these three different settings. Bioinformatic analysis highlighted key sequence types (STs), AMR gene and plasmid profiles in the three datasets. These techniques and daily sampling helped to characterise the dynamic nature of E. coli colonisation in travellers to Laos, showing the high level of MDR E. coli present and the rapid acquisition and loss of STs during their traveller stay. Detailed characterisation of the plasmids carried by these MDR E. coli highlighted the diversity of plasmids and the diversity of AMR gene settings within these AMR plasmids. Study of E. coli in ICU patients, demonstrated the presence of persistent colonisation during a hospital stay, and the lack of transmission, AMR carriage and ST diversity, widely differing from the Laos traveller dataset. Strain level analysis revealed circulating strains in the traveller population and lack of circulating strains in the ICU population. The clinical infection E. coli isolated from neonates with meningitis or bloodstream infection (BSI), demonstrated that over 47-years resistance carriage remained surprisingly low but there were interesting changes to the composition of the ST population causing invasive disease, including the loss of a predominant ST, ST567, which had not been previously widely reported. Overall, this study revealed the different dynamics of the three E. coli populations investigated. Strain level sampling in these relevant settings showed the three very different antimicrobial resistance profiles and different ST populations in the absence of any antibiotic selection

Multidrug resistance plasmids commonly reprogram the expression of metabolic genes in <i>Escherichia coli</i>

Multidrug-resistant Escherichia coli is a leading cause of global mortality. Transfer of plasmids carrying genes encoding beta-lactamases, carbapenamases, and colistin resistance between lineages is driving the rising rates of hard-to-treat nosocomial and community infections. Multidrug resistance (MDR) plasmid acquisition commonly causes transcriptional disruption, and while a number of studies have shown strain-specific fitness and transcriptional effects of an MDR plasmid across diverse bacterial lineages, fewer studies have compared the impacts of different MDR plasmids in a common bacterial host. As such, our ability to predict which MDR plasmids are the most likely to be maintained and spread in bacterial populations is limited. Here, we introduced eight diverse MDR plasmids encoding resistances against a range of clinically important antibiotics into E. coli K-12 MG1655 and measured their fitness costs and transcriptional impacts. The scale of the transcriptional responses varied substantially between plasmids, ranging from &gt;650 to &lt;20 chromosomal genes being differentially expressed. However, the scale of regulatory disruption did not correlate significantly with the magnitude of the plasmid fitness cost, which also varied between plasmids. The identities of differentially expressed genes differed between transconjugants, although the expression of certain metabolic genes and functions were convergently affected by multiple plasmids, including the downregulation of genes involved in L-methionine transport and metabolism. Our data show the complexity of the interaction between host genetic background and plasmid genetic background in determining the impact of MDR plasmid acquisition on E. coli.IMPORTANCE: The increase in infections that are resistant to multiple classes of antibiotics, including those isolates that carry carbapenamases, beta-lactamases, and colistin resistance genes, is of global concern. Many of these resistances are spread by conjugative plasmids. Understanding more about how an isolate responds to an incoming plasmid that encodes antibiotic resistance will provide information that could be used to predict the emergence of MDR lineages. Here, the identification of metabolic networks as being particularly sensitive to incoming plasmids suggests the possible targets for reducing plasmid transfer. </p

Effectiveness and cost-effectiveness of community singing on the mental health related quality of life of the older population: a randomized controlled trial

Background: As the population ages, older people account for a greater proportion of the health and social care budget. While some research has been conducted on the use of music therapy for specific clinical populations, little rigorous research has been conducted looking at the value of community singing on the mental health-related quality of life of older people. Aims: To evaluate the effectiveness and cost-effectiveness of community group singing for a population of older people in England. Method: A pilot pragmatic individual randomized controlled trial comparing group singing versus usual activities in those aged 60 years or more. Results: 258 participants were recruited across 5 centres in East Kent. At 6 months post-randomisation significant differences were observed in terms of mental health related quality of life measured using the SF12, mean difference 2.35 (95% CI 0.06 to 4.76) in favour of group singing. In addition the intervention was found to be marginally more cost-effective than usual activities. At 3 months significant differences were observed in terms of mental health components of quality of life (4.77; 2.53 to 7.01) anxiety (-1.78; -2.5; -1.06) and depression (-1.52; -2.13 to -0.92). Conclusions: Community group singing appears to have a significant effect on mental health related quality of life, anxiety and depression and may be a useful intervention to maintain and enhance the mental health of the older population

Integration of vanHAX downstream of a ribosomal RNA operon restores vancomycin resistance in a susceptible Enterococcus faecium strain

During the genomic characterisation of Enterococcus faecium strains (n = 39) collected in a haematology ward, we identified an isolate (OI25), which contained vanA-type vancomycin resistance genes but was phenotypically susceptible to vancomycin. OI25 could revert to resistance when cultured in the presence of vancomycin and was thus considered to be vancomycin-variable. Long-read sequencing was used to identify structural variations within the vancomycin resistance region of OI25 and to uncover its resistance reversion mechanism. We found that OI25 has a reduced ability to positively regulate expression of the vanHAX genes in the presence of vancomycin, which was associated with the insertion of an IS6-family element within the promoter region and the first 50 bp of the vanR gene. The vancomycin-resistant revertant isolates constitutively expressed vanHAX genes at levels up to 36,000-fold greater than OI25 via co-transcription with a ribosomal RNA operon. The vancomycin-resistant revertants did not exhibit a significant growth defect. During VRE outbreaks, attention should be paid to contemporaneous vancomycin-susceptible strains as these may carry silent vancomycin resistance genes that can be activated through genomic rearrangements

Real-time sampling of travelers shows intestinal colonization by multidrug-resistant bacteria to be a dynamic process with multiple transient acquisitions

AbstractBackgroundAntimicrobial resistance (AMR) is highly prevalent in low- and middle-income countries (LMIC). International travel contributes substantially to the global spread of intestinal multidrug-resistant gram-negative (MDR-GN) bacteria. Of the 100 million annual visitors to LMIC, 30–70% become colonized by MDR-GN bacteria. The phenomenon has been well documented, but since sampling has only been conducted after travelers’ return home, data on the actual colonization process are scarce. We aimed to characterize colonization dynamics by exploring stool samples abroad on a daily basis while visiting LMIC.MethodsA group of 20 European volunteers visiting Lao People’s Democratic Republic for three weeks provided daily stool samples and filled in daily questionnaires. Acquisition of extended-spectrum beta-lactamase-producing gram-negative bacteria (ESBL-GN) was examined by selective stool cultures followed by whole-genome sequencing (WGS) of isolates.ResultsWhile colonization rates were 70% at the end of the study, daily sampling revealed that all participants had acquired ESBL-GN at some time point during their overseas stay, the colonization status varying day by day. WGS analysis ascribed the transient pattern of colonization to sequential acquisition of new strains, resulting in a loss of detectable colonization by the initial MDR-GN strains. All but one participant acquired multiple strains (n=2–7). Of the total of 83 unique strains identified (53 E. coli, 10 Klebsiella, 20 other ESBL-GN species), some were shared by as many as four subjects.ConclusionsThis is the first study to characterize in real time the dynamics of acquiring MDR-GN during travel. Our data show multiple transient colonization events indicative of constant microbial competition.</jats:sec

Non-antibiotic pharmaceuticals are toxic against <i>Escherichia coli</i> with no evolution of cross-resistance to antibiotics

Antimicrobial resistance can arise in the natural environment via prolonged exposure to the effluent released by manufacturing facilities. In addition to antibiotics, pharmaceutical plants also produce non-antibiotic pharmaceuticals, both the active ingredients and other components of the formulations. The effect of these on the surrounding microbial communities is less clear. We aimed to assess whether non-antibiotic pharmaceuticals and other compounds produced by pharmaceutical plants have inherent toxicity, and whether long-term exposure might result in significant genetic changes or select for cross-resistance to antibiotics. To this end, we screened four non-antibiotic pharmaceuticals (acetaminophen, ibuprofen, propranolol, metformin) and titanium dioxide for toxicity against Escherichia coli K-12 MG1655 and conducted a 30 day selection experiment to assess the effect of long-term exposure. All compounds reduced the maximum optical density reached by E. coli at a range of concentrations including one of environmental relevance, with transcriptome analysis identifying upregulated genes related to stress response and multidrug efflux in response ibuprofen treatment. The compounds did not select for significant genetic changes following a 30 day exposure, and no evidence of selection for cross-resistance to antibiotics was observed for population evolved in the presence of ibuprofen in spite of the differential gene expression after exposure to this compound. This work suggests that these compounds, at environmental concentrations, do not select for cross-resistance to antibiotics in E. coli

Strategies for assessing renal function prior to outpatient contrast-enhanced CT: a UK survey

YesThe purpose of this paper is to identify current UK screening practices prior to contrast-enhanced CT. To determine the patient management strategies to minimize the risk of contrast-induced acute kidney injury (CI-AKI) risk in outpatients. An invitation to complete an electronic survey was distributed to the CT managers of 174 UK adult National Health Service hospital trusts. The survey included questions related to local protocols and national guidance on which these are based. Details of the assessment of renal function prior to imaging and thresholds for contrast contraindication and patient management were also sought. A response rate of 47.1% was received. Almost all sites had a policy in place for contrast administration (n = 80/82; 97.6%). The majority of sites require a blood test on outpatients undergoing a contrast-enhanced CT scan (n = 75/82; 91.5%); however, some (15/75; 20.0%) sites only check the result in patients at high risk and a small number (7/82; 8.5%) of sites indicated that it was a referrer responsibility. The estimated glomerular filtration rate (eGFR) or serum creatinine (SCr) result threshold at which i.v. contrast was contraindicated varied and 19 different threshold levels of eGFR or SCr were identified, each leading to different prophylactic strategies. Inconsistency was noted in the provision of follow-up blood tests after contrast administration. The wide variation in practice reflects inconsistencies in published guidance. Evidence-based consensuses of which patients to test and subsequent risk thresholds will aid clinicians identify those patients in which the risk of CI-AKI is clinically significant but manageable. There is also a need to determine the value of the various prophylactic strategies, follow-up regimen and efficient service delivery pathways. This survey has identified that further work is required to define which patients are high risk, confirm those which require renal function testing prior to contrast administration and how best to manage patients at risk of CI-AKI. The role of new technologies within this service delivery pathway requires further investigation

The effectiveness and cost-effectiveness of a participative community singing programme as a health promotion initiative for older people: protocol for a randomised controlled trial

Background The growth in numbers of older people represents a considerable cost to health and social care services in the United Kingdom. There is an acknowledged need to address issues of social exclusion and both the physicial and mental health of this age group. In recent years there has been much interest in the potential contribution of the arts to the health of communities and individuals. There is some evidence that participative singing may be of benefit to older people, however studies to date are limited in number and have lacked rigour. There is therefore a need to build on this knowledge base to provide more quantifiable evidence of both effectiveness and cost efffectness of singing as a health intervention of this population group. Methods The proposed study is a pragmatic randomised controlled trial with two parallel arms. The primary hypothesis is that singing groups for older people improve both physical and mental aspects of qualaity of life when compared to usual activities. Potential participants will be volunteers over 60 years living in the community and recruited through publicity. Eligible and consenting participants will be randomized to either a singing group or a control group. Singing groups will take part in a twelve week planned programme of singing and control groups will continue with usual activities. The primary outcome measure will be the York SF-12, a health related quality of life measure which will be administered at baseline, three and six months after baseline. The study will evaluate both effectiveness and cost-effectiveness. Discussion This study proposes to add to the exisiting body of evidence on the value of singing for older people by using a rigorous methodological design, which includes a power calucation, a standardised intervention and assessment of cost-effectiveness. It should be regarded as a stage in a progressive programme of studies in this area. If group singing is found to be effective and cost-effective it may offer an alternative means to maintaining the health of people in later life

Inhibition of Intestinal Bile Acid Transporter Slc10a2 Improves Triglyceride Metabolism and Normalizes Elevated Plasma Glucose Levels in Mice

Interruption of the enterohepatic circulation of bile acids increases cholesterol catabolism, thereby stimulating hepatic cholesterol synthesis from acetate. We hypothesized that such treatment should lower the hepatic acetate pool which may alter triglyceride and glucose metabolism. We explored this using mice deficient of the ileal sodium-dependent BA transporter (Slc10a2) and ob/ob mice treated with a specific inhibitor of Slc10a2. Plasma TG levels were reduced in Slc10a2-deficient mice, and when challenged with a sucrose-rich diet, they displayed a reduced response in hepatic TG production as observed from the mRNA levels of several key enzymes in fatty acid synthesis. This effect was paralleled by a diminished induction of mature sterol regulatory element-binding protein 1c (Srebp1c). Unexpectedly, the SR-diet induced intestinal fibroblast growth factor (FGF) 15 mRNA and normalized bile acid synthesis in Slc10a2−/− mice. Pharmacologic inhibition of Slc10a2 in diabetic ob/ob mice reduced serum glucose, insulin and TGs, as well as hepatic mRNA levels of Srebp1c and its target genes. These responses are contrary to those reported following treatment of mice with a bile acid binding resin. Moreover, when key metabolic signal transduction pathways in the liver were investigated, those of Mek1/2 - Erk1/2 and Akt were blunted after treatment of ob/ob mice with the Slc10a2 inhibitor. It is concluded that abrogation of Slc10a2 reduces hepatic Srebp1c activity and serum TGs, and in the diabetic ob/ob model it also reduces glucose and insulin levels. Hence, targeting of Slc10a2 may be a promising strategy to treat hypertriglyceridemia and diabetes