The growing crisis of antibiotic resistance is a major threat to ecosystems and human health. Infections caused by known and emerging antibiotic resistant pathogens are on the rise globally, with approximately 700,000 deaths per year caused by antibiotic resistant bacteria (1). In the United States, infections from antibiotic resistant bacteria cause more than 2 million illnesses and 23,000 deaths (2). Antibiotic resistant bacteria and antibiotic resistance genes are released into aquatic ecosystems through hospital waste, residential sewer lines and animal agricultural waste streams. Animal agriculture accounts for approximately 70% of antibiotic use in the United States (3). In agricultural ecosystems, runoff, land-applied fertilizer and waste lagoons can all contribute to the spread of antibiotic resistance. In urban ecosystems, sewage spills and other wastewater inputs contribute to the spread of antibiotic resistance. Environmental matrices, such as soil and water, can provide habitat, serving as reservoirs to potentially promote the spread of resistance. Research addressing antibiotic resistance primarily focuses on monitoring clinical occurrence and nosocomial infections (acquired in hospitals),but the natural environment also plays a role in the spread of antibiotic resistance. The consequences to aquatic ecosystems are not often studied and not well understood. Antibiotic resistance genes can transfer between bacteria through transduction, transformation and conjugation, potentially persisting in non-pathogenic environmental bacteria. Environmental reservoirs of antibiotics, antibiotic resistant bacteria and antibiotic resistance genes should be considered and integrated into frameworks to improve monitoring, regulation and management of urban and rural watersheds. The research presented in this doctoral dissertation includes field and laboratory studies designed to assess the prevalence and persistence of antibiotic resistant bacteria and antibiotic resistance genes in aquatic environments, with a focus on vancomycin-resistant enterococci, which are considered a major threat in the United States and top priority pathogens according to the Centers for Disease Control (2). The vanA gene associated with high-level resistance is located on mobile plasmids and associated with clinical infections, predominantly in the species Enterococcus faecium. E. faecium can cause bacteremia, endocarditis, pelvic infections and more (4). When vancomycin, often the last line of treatment for these infections, is no longer effective, the health burdens increase both financially and physically and infections can be fatal. Chapter 1 summarizes background and review of antibiotic resistance in the environment, including a co-authored review of culture-based methods to detect antibiotic resistant bacteria and antibiotic resistance genes in in the environment (previously published in the Journal of Environmental Quality (5). In Chapter 2, a field study was performed to investigate the occurrence and persistence of vancomycin-resistant enterococci and vanA in a sewage spill in Pinellas County, Florida, previously published in the journal Applied and Environmental Microbiology (6). In Chapter 3, antibiotic resistance genes were quantified to study their persistence in poultry litter microcosms (manuscript in prep). In Chapter 4, microcosms were used to assess how nutrients and plasmid-associated vancomycin resistance affect survival among E. faecium strains (in process of submitting for publication at Applied and Environmental Microbiology). Antibiotic resistance is a public health crisis and the results of the studies presented here contribute data towards a better understanding of environmental reservoirs of antibiotic resistant bacteria and antibiotic resistance genes. The research has broad implications for public health, environmental policy and ecosystem management.

Dans le contexte de l'antibiorésistance, l'objet de ce doctorat vise à évaluer l'impact d'antibiotiques sur le microbiote intestinal de porcelets. La colistine et le ceftiofur, pour lesquels les résistances incluent essentiellement et respectivement mutations chromosomiques et gènes plasmidiques, ont été utilisés. La colistine a significativement réduit la population des entérobactéries, mais aucun E. coli résistant n'a été détecté. L'administration de ceftiofur a eu un impact limité sur les populations bactériennes composant l'écosystème digestif mais a conduit à une forte sélection et à la diffusion d'un gène plasmidique codant pour une bêta-lactamase à spectre étendu. Puis, dans le cadre de la réglementation visant à diminuer l'expérimentation animale, un modèle in vitro colique porcin, nommé PigutIVM, a été mis au point afin de simuler l'environnement digestif du porcelet et a permis de confirmer, in vitro, l'effet observé in vivo de la colistine sur le microbiote. Cet outil a ensuite été utilisé pour évaluer l'impact d'un probiotique, Saccharomyces cerevisiae, comme alternative aux antibiotiques. Le modèle PigutIVM devrait se positionner comme un outil de prédiction pertinent dans les domaines d'investigation aussi bien nutritionnels que pharmacologiques. / In the context of antibiotic resistance, the aim of the current PhD work is to assess the impact of antibiotics on intestinal microbiota of piglets. Two antibiotics i.e. colistin and ceftiofur, for which the main resistances include respectively chromosomal mutations and plasmid genes have been used. Colistin significantly reduced the population of Enterobacteriaceae, but there was no selection of resistant E. coli. The administration of ceftiofur had a limited impact on the bacterial populations that make up the digestive ecosystem but it led to strong selection and dissemination of a plasmid gene encoding an extended-spectrum beta-lactamase. Then, in the framework of regulations to reduce animal testing, an in vitro model of colonic pig named PigutIVM was developed in order to simulate the digestive environment of the piglet and then check the effect of colistin on the microbiota simulated in PigutIVM in vitro. Therefore both the approaches i.e. in vivo and in vitro were compared in order to check the effect of colistin on intestinal microbiota of piglets. This tool was then used to evaluate the impact of a probiotic i.e. Saccharomyces cerevisiae, as alternative to antibiotics. Therefore we assume that this PigutIVM model should be positioned as a relevant predictive tool in the fields of nutritional and pharmacological investigations.

Worldwide there is a growing concern of the emergence and evolution of antimicrobial resistance among bacterial pathogens, which poses a threat to human and animal health. The extensive use and misuse of antimicrobials in human and veterinary clinical therapy and agricultural practices have been a major selective force for the emergence, selection, and dissemination of antimicrobial resistant bacteria and resistant genes. Commensal bacteria constitute a reservoir of resistant genes and their level of resistance is considered to be a good indicator for resistance problems to be expected in pathogens. The monitoring of the prevalence of resistance in indicator bacteria such as faecal Escherichia coli and enterococci in different human and animal populations allows the comparison of the prevalence of resistance and to detect transfer between animals and humans and vice versa. Antimicrobial resistance has however, been found in the bacteria of wildlife not exposed to antimicrobials and living in remote areas of this earth. This has implications for resistance control strategies. Previous studies on antimicrobial resistance in wildlife have yielded contrasting results, such as an almost complete absence of resistance in enterobacteria isolated from moose, deer and vole in Finland compared to a high prevalence of resistance in faecal bacteria from wild rodents living in northwest England, which are possibly due to differences in the ecological systems and the proximity to anthropogenic activities. This study further investigates the phenomenon of antimicrobial resistance in wildlife. A previous study conducted in the conservancy area of Kruger National Park (KNP) within South Africa showed that surface water could be a possible source of antimicrobial resistance in unexposed animal populations and that impala (Aepyceros melampus) were good sentinel animals for the documentation of antimicrobial resistance through rivers. This current study followed on this hypothesis and investigated the prevalence of resistance in commensal bacteria isolated from impala and their water sources in KNP. The following four perennial river systems were selected: the Olifants, the Letaba, the Crocodile, the Sabie-Sand Rivers. Samples of river water (n=11) and faeces (n=165) were collected at 11 different sites along these rivers. Samples were directly plated and resistant colonies were selected by means of discs containing antimicrobials (direct plating method). Resistant colonies that grew in the presence of antimicrobials were cultured and identified. Isolates of E. coli (n=12), Enterobacter cloacae (n=49), Pantoea species (n=9), Enterococcus faecalis (n=59), Enterococcus faecium (n=4) and Enterococcus durans (n=64) were tested for susceptibility to a selection of commonly used veterinary antimicrobial drugs. Susceptibility to 18 antimicrobial drugs was determined by means of minimum inhibitory concentrations (MIC) using a commercial MIC test (Sensititre® Bovine/Porcine plate format BOP06F). Our results allow us to give further support to our working hypothesis that antimicrobial resistance, as evidenced in the impala faeces, may have been due to the impala drinking from the polluted rivers, knowing full well that impala are not routinely subjected to any form of antimicrobial treatment. Although the isolates obtained from the water sources were not as many as those obtained from the faecal samples, a degree of resistance was also observed across all the four river systems that we isolated bacteria from, and this was also evident in the faecal samples as well. Our results also further add to the importance of wildlife as sentinels in environmental antimicrobial resistance studies. / Dissertation (MSc)--University of Pretoria, 2013. / gm2014 / Veterinary Tropical Diseases / Unrestricted

Hajj, the annual Islamic pilgrimage to Makkah, Saudi Arabia, is a unique mass gathering event that brings more than 2 million individuals from around the world. Several public health considerations, such as the spread of infectious diseases, must be taken into account with this large temporary influx of people. Gastrointestinal diseases, such as diarrhea, are common at Hajj, yet little is known about the etiology. The human gut microbiome, collection of organisms residing within the intestinal tract, has been under intense study recently, since next generation DNA sequencing technologies allow for extensive surveying of genetic material found in complex biological samples, such as those containing many different organisms. Thus, using 16S rRNA and metagenomic shotgun sequencing, we have characterized the gut microbiome of over 612 pilgrims with and without diarrhea. Several metadata factors, such as hospitalization and different comorbidities, were found to have significant effects on the overall gut microbiome composition. Metagenomic shotgun sequencing efforts revealed the presence of antimicrobial resistance genes originating from disparate regions from around the world. This study provides a snapshot of information concerning the health status of the gut microbiome of Hajj pilgrims and provides more context to the investigation of how to best prepare for mass gathering events.

In healthcare settings and beyond, the ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) among other pathogens exchange antibiotic resistance and virulence factors and emerge as new infectious clones. According to the Saudi General Authority for Statistics (stats.gov.sa), Saudi Arabia is a country where more than 27 million pilgrims meet in annual continual mass-gathering events. This massive influx of people could introduce novel pathogens to the community that could not necessarily be detected with traditional culture-dependent clinical microbiological tests. Conventional clinical microbiology and environmental pathogen detection methods have had many limitations and narrow search scope. These methods can only target known and culturable pathogens. Over the past decade, applications of next-generation sequencing (NGS) and bioinformatics tools have revolutionized the way pathogens are detected and their relevant phenotypes such as clonal types, antibiotic resistance are predicted to aid in clinical decision making as additional practice to traditional clinical microbiology-based testing protocols. The aim of this study was to apply whole-genome sequencing (WGS) and bioinformatic analysis tools on clinical samples and bacterial isolates in order to pave the way for transforming current clinical microbiology practices in a tertiary referral hospital in Jeddah, Saudi Arabia. My attempt to utilize WGS as a tool on pathogenic strains in this study combined with the clinical data has resulted in discovering a silent outbreak of an emerging hypervirulent strain of Klebsiella pneumoniae (Chapter 2). Analysis of the strains antimicrobial profiles genetically has yielded the first characterization of a misidentified Klebsiella quasipneumoniae harboring plasmid-mediated carbapenemases of Klebsiella pneumoniae carbapenemases (KPC) (Chapter 3). Similarly, I was able to study mobile colistin resistance genes in the isolates and identify a novel occurrence of mcr-1 and mcr-8 (Chapter 4). I applied clinical metagenomic protocol on an intestinal biopsy of an inflammatory bowel disease patient with Crohn’s disease, where I identified an association of three co-occurring and an actively replicating non-tuberculosis mycobacteria (Chapter 5). The deployment of whole-genome sequencing and metagenomic in infectious disease surveillance and diagnostics could prove beneficial in limiting epidemics and detect transmission patterns of antimicrobial-resistant genes.

京都大学 / 0048 / 新制・課程博士 / 博士(工学) / 甲第22762号 / 工博第4761号 / 新制||工||1745(附属図書館) / 京都大学大学院工学研究科都市環境工学専攻 / (主査)教授 田中 宏明, 教授 米田 稔, 准教授 松田 知成 / 学位規則第4条第1項該当 / Doctor of Philosophy (Engineering) / Kyoto University / DFAM

Background The antimicrobial stewardship is developed to provide a guide on the responsible use of antimicrobial drugs. Thus, slowing down the development of antimicrobial resistance. However, the nurse's role in antimicrobial stewardship is not clarified. Failure toinclude the nurse within the antimicrobial stewardship guidelines may result in poor execution of antimicrobial stewardship.Aim To explore the role of nurses in antimicrobial stewardship and how it can be practically implemented within the medical field.Method This is a literature review where seven qualitative studies, two quantitative studies and a mix-methods study examines the nurse's role in antimicrobial stewardship.Results Two main themes and five sub-themes were created. The two main themes were clinical role and collaboration. The clinical role described the nurse's role as a patient advocate and the nurse's contribution to antimicrobial stewardship through monitoring and evaluation of the patient and treatment, as well as through safe sampling, drug administration and hygiene. The collaboration showed and identified the nurse's role as a communicator and educator. Conclusion Conclusions that can be drawn from the literature review are that the potential roles the nurse may have in antimicrobial stewardship are many and those we have identified are already included in the nurse's daily work.

<div>Pulsed electric fields (PEFs) can induce numerous biophysical phenomena, especially perturbation of the outer and inner membranes, that may be used for applications that include nonthermal pasteurization, enhanced permeabilization of tumors to improve the transport of chemotherapeutics for cancer therapy, and enhanced membrane permeabilization of individual cells to enhance RNA and DNA delivery for gene therapy. The applied electric field and pulse duration determine the density, size, and reversibility of the created membrane pores. PEFs with durations longer than the outer membrane’s charging time will induce pore formation with the potential for application in irreversible electroporation for cancer therapy and microorganism inactivation. Shorter duration PEFs, particularly on the nanosecond timescale (nsPEFs), induce a larger density of smaller membrane pores with the potential to permeabilize intracellular membranes, such as the mitochondria, to induce programmed cell death. Thus, the PEFs can effectively kill multiple types of cells, dependent upon the cells. This thesis assesses the ability of nsPEFs to kill different cell types, specifically microorganisms with and without antibiotics as well as varying the parameters to affect populations of immortalized leukemia cells (Jurkats).</div><div>Antibiotic resistance has been an acknowledged challenge since the initial development of penicillin; however, recent discoveries by the CDC and the WHO of microorganisms resistant to last line of defense drugs combined with predictions of potential infection cases reaching 50 million a year globally and the absence new drugs in the discovery pipeline highlight the need to develop novel ways to combat and overcome these resistance mechanisms. Repurposing drugs, exploring nature for new drugs, and developing enzymes to counter the resistance mechanisms may provide potential alternatives for addressing the scarcity of antibiotics effective against gram-negative infections. One may also leverage the abundance of drugs effective against gram-positive infections by using nsPEFs to make them effective against gram-negative infections, including bacterial species with multiple natural and acquired resistance mechanisms. Numerous drug and microbial combinations for different doses and pulse treatments were tested and presented here.</div><div>Low intensity PEFs may selectively target cell populations at different stages of the cell cycle (quiescence and mitosis) to modify cancer cell population dynamics. Experimental studies of cancer cell growth when exposed to a low number of nsPEFs, while varying pulse duration, field intensity and number of pulses reveals a threshold beyond which cell recovery is not possible, but also a point of diminishing returns if cell death is the intention. A theory comprised of coupled differential equations representing the proliferating and quiescent cells showed how changing PEF parameters altered the behavior of these cell populations after treatment. These results may provide important information on the impact of PEFs with sub-threshold intensities and durations on cell population growth and potential recurrence.</div>

The purpose of this research is to support the California Department of Food & Agriculture (CDFA) with education and outreach about recent changes regarding antimicrobial use in livestock, and to enhance their emergency communications network. This was done by characterizing a population of small-scale livestock owners underserved with regards to educational outreach about animal health issues, such as antimicrobial use rules. The project also seeks to enhance public understanding of the importance of responsible antimicrobial use in animal and human health. To accomplish this, the study used a survey administered in person at local farm supply stores and online to investigate the level of understanding of antimicrobial rules among livestock owners in San Luis Obispo county. The survey gathered information about the livestock owners’ practices, connection to livestock groups, and access to information pertaining to animal health among other things. The results of the survey showed that respondents were largely unaware of new rules pertaining to use of antimicrobials in livestock. Familiarity with California rules of this kind was used as an indicator of access to information about animal health and was found to correlate positively with knowledge about antimicrobial resistance and familiarity with federal rules concerning antimicrobial use in livestock. As predicted, respondents with a connection to 4-H and FFA had greater familiarity with both federal and state rules concerning antimicrobial use in livestock.

No description available.