The reduced efficacy of antibiotics in treating common infections is one of the most pressing health concerns of the 21st Century. Increasing evidence links the widespread use of antibiotics in livestock production to the transfer of bacteria carrying antibiotic resistance genes to the broader environment. It is therefore critical to understand the persistence and dissemination of resistance in agricultural soils to understand potential threats to consumers. The goal of this large-scale agricultural field experiment was to identify the effects of crop (lettuce, radish) and fertilizer type (inorganic, compost, raw manure) on the incidence and persistence of antibiotic-resistant fecal coliforms, a common family of fecal indicator bacteria used to track the environmental spread of antibiotic resistance. Soil samples were collected eight times over a 120-day period and analyzed for fecal coliforms utlizing a suite of MacConkey agars supplemented with different antibiotics (ceftazidime, clindamycin, erythromycin, sulfamethoxazole, and tetracycline). Given the number of samples with resistant fecal coliform concentrations below the limit of detection, analyses to identify the effects of soil treatment and crop relied on Zero-inflated Poisson Regressions. Antibiotic-resistant culturable fecal coliforms were recoverable from soils across all treatments immediately following application, though persistence throughout the experiment varied by antibiotic. Sulfamethoxazole- and tetracycline-resistant fecal coliforms were nondetectable after Day 1; this was expected, as the cattle supplying the manure amendments were not treated with these antibiotics or similar analogs. Clindamycin- and erythromycin-resistant fecal coliforms were nondetectable after 42 days but rebounded on Day 90 in the soil; both of these drugs were of the same antibiotic class as the ones used to treat the dairy cattle during the manure collection period. Ceftazidime-resistant fecal coliform levels were consistently high throughout the duration of the growing season. No statistical differences were observed between root and aboveground crops. Results suggest that soils amended with raw or composted dairy manure are at risk of contamination with antibiotic resistant fecal coliforms; however, composting decreased the antibiotic resistant fecal coliform levels of the macrolide (erythromycin) and lincosamide (clindamycin) antibiotic classes administered to the dairy cattle (cephapirin and pirlimycin). / Master of Science / The reduced efficacy of antibiotics in treating common infections is one of the most pressing health concerns of the 21st Century. Increasing evidence links the widespread use of antibiotics in livestock production to the transfer of bacteria carrying antibiotic resistance genes to the broader environment. It is therefore critical to understand the persistence and dissemination of resistance in agricultural soils to understand potential threats to consumers. The goal of this large-scale agricultural field experiment was to identify the effects of crop (lettuce, radish) and fertilizer type (inorganic, compost, raw manure) on the incidence and persistence of antibiotic-resistant fecal coliforms, a common family of fecal indicator bacteria used to track the environmental spread of antibiotic resistance. Over the course of 120 days, samples were collected from field plots to identify if there were antibiotic-resistant bacteria (ARBs) in the soil. This study was partially motivated as a means to evaluate the Federal Drug Administration’s Food Modernization Safety Act updated manure treatment guidelines in decreasing potential pathogenic bacteria in soils used to grow vegetables for human consumption. Antibiotic-resistant bacteria were recoverable from soils across all fertilizer types immediately following application, though persistence throughout the experiment varied by antibiotic tested. From the above findings, compost amended soils had greater quantities of total and ceftazidime-resistant bacteria. However, composting did show a significant decrease in the antibiotic-resistant bacteria levels found in the same antibiotic classes, macrolides and lancosimides, that the dairy cattle were treated with at the beginning of this study.

Antibiotic resistance is a growing human health threat, making infections more difficult to treat and increasing fatalities from and cost of treatment of associated diseases. The rise of multidrug resistant pathogens threatens a return to the pre-antibiotic era where even the most common infections may be impossible to treat. It is estimated that the majority of global antibiotic use, and use in the U.S., is dedicated towards livestock, where they are used to promote growth, treat, or prevent disease. Given that exposure to antibiotics selects for antibiotic resistant bacteria (ARBs) and can stimulate the horizontal transfer of their associated antibiotic resistance genes (ARGs), it is important to examine livestock operations as a reservoir of resistance. Correspondingly, there is growing interest in identifying how agricultural practices can limit the potential for spread of antibiotic resistance through the "farm to fork continuum," starting with antibiotic use practices, manure management and land application and ending with the spread of ARBs and ARGs present onto edible crops and serving as a route of exposure to consumers. This study focused specifically on the effect of composting on the prevalence of ARBs and ARGs in cattle manure. Three composting trials were performed: small-scale, heat-controlled, and large-scale. The small-scale composting trial compared dairy and beef manures, with or without antibiotic treatment (treated beef cattle received chlortetracycline, sulfamethazine, and tylosin while treated dairy cattle received cephapirin and pirlimycin), subject to either static or turned composting. The heat-controlled composting trial examined only dairy manure, with or without antibiotic treatment, subject to static composting, but using external heat tape applied to the composting tumblers to extend the duration of the thermophilic (>55°C) temperature range. The large-scale composting trial examined dairy manure, with or without antibiotic treatment, subject to static composting at a much larger scale that is more realistic to typical farm practices. Samples were analyzed to assess phenotypic resistance using the Kirby Bauer disk diffusion method and by diluting and plating onto antibiotic-supplemented agar. Genetic markers of resistance were also assessed using quantitative polymerase chain reaction (qPCR) to quantify sul1 and tet(W) ARGs; metagenomic DNA sequencing and analysis were also performed to assess and compare total ARG abundance and types across all samples. Results indicate that composting can enrich indicators of phenotypic and genetic resistance traits to certain antibiotics, but that most ARGs are successfully attenuated during composting, as evidenced by the metagenomic sequencing. Maintaining thermophilic composting temperatures for adequate time is necessary for the effective elimination of enteric bacteria. This study suggests that indicator bacteria that survive composting tend to be more resistant than those in the original raw manure; however, extending the thermophilic stage of composting, as was done in the heat-controlled trial, can reduce target indicator bacteria below detection limits. Of the two ARGs specifically quantified via qPCR, prior administration of antibiotics to cattle only had a significant impact on tet(W). There was not an obvious difference in the final antibiotic resistance profiles in the finished beef versus dairy manure composts according to metagenomics analysis. Based on these results, composting is promising as a method of attenuating ARGs, but further research is necessary to examine in depth all of the complex interactions that occur during the composting process to maximize performance. If not applied appropriately, e.g., if time and temperature guidelines are not enforced, then there is potential that composting could exacerbate the spread of certain types of antibiotic resistance. / Master of Science / Antibiotics are drugs that are used to treat bacterial infections by killing the bacteria that cause the infection. Bacterial infections now exist that are resistant to several antibiotics; which are extremely difficult and costly to treat. Many antibiotics are used in the agriculture industry where they are used to promote growth, treat, or prevent disease in livestock animals. The antibiotics may then cause an increase in antibiotic resistance in bacteria by encouraging changes to the DNA of the bacteria which allow them to survive in the presence of antibiotics that would normally kill them. These DNA segments are called antibiotic resistance genes. Once developed, bacteria can share resistance genes among themselves, allowing for single bacteria that can resist several types of antibiotics. For this reason, it is important to see if it is possible to prevent the spread of antibiotic resistance from animal agriculture to people. One way that people could be affected would be if produce were exposed to resistant bacteria when grown in soil that had been fertilized with manure or compost. This study looks at the impact of composting on the presence and amount of antibiotic resistance genes in composted cattle manure. Three composting trials were performed: small-scale, heat-controlled, and large-scale. The small-scale composting trial compared dairy and beef manures, with or without antibiotic treatment, with or without regular turning during composting. The heat-controlled composting trial examined only dairy manure, with or without antibiotic treatment, without regular turning during composting, but using external heat to maintain high temperatures. The large-scale composting trial examined dairy manure, with or without antibiotic treatment, without regular turning during composting, but at a larger scale that is more realistic to how composting is actually performed on farms. Antibiotic resistance of compost bacteria was tested by growing bacteria on nutrient-dense plates containing antibiotic disks and measuring how much each antibiotic prevented the growth of the bacteria, in terms of the diameter about each disk where bacteria did not grow. Individual target resistance genes were measured throughout the study by using a method called qPCR. Metagenomic analysis was performed to identify all of the genes, especially resistance genes, in each of the samples. Results v show that composting may increase antibiotic resistance in bacteria that survive the composting process, but that most resistance genes are themselves reduced. The key to successful composting is maintaining high temperatures for as long as possible; this is necessary to kill off infectious bacteria. Extending the high temperature (>55°C) phase of composting is a potential method for improving the effectiveness of composting in eliminating pathogens and destroying resistance genes. Results were not significantly affected by whether antibiotics were given to the cattle and were not different between dairy or beef cattle. Based on these results, composting is a promising method of reducing resistance genes in composted manure, but further research is necessary to maximize performance. If not performed correctly, composting could have the opposite effect and be detrimental.

京都大学 / 新制・課程博士 / 博士(工学) / 甲第25270号 / 工博第5229号 / 新制||工||1997(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 西村 文武, 教授 松田 知成, 准教授 浅田 安廣, 准教授 井原 賢(高知大学) / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DGAM

Antibiotic-resistant bacteria(ARB) is causing increased health risk and cost to society. Mathematical models have been developed to study the transmission of resistant bacteria and the efficacy of preventive measures to slow its spread within a hospital setting. The majority of these models have assumed a constant total hospital population with the admission and discharge rates being equal throughout the duration. But a typical hospital population varies from day to day and season to season. In this thesis, we apply variable admission and discharge daily rates to existing deterministic and stochastic models which examine the transmission of single and dual resistant bacteria. We perform stability and equilibrium analyses as well as a sensitivity analysis on the resulting model..

Smittsamma sjukdomar kostar det svenska samhället enorma summor varje år. Behandlingen av smittade patienter har tidigare uppskattats till 5-10 miljarder svenska kronor årligen. Vidare estimeras de förebyggande åtgärderna kosta samhället runt en miljard svenska kronor. Detta betyder att det finns en ekonomisk drivkraft för att reducera antalet smittade patienter inom vården, speciellt de fall som är orsakade av resistenta bakterier. Samtidigt pågår det en debatt om resistenta bakterier och antibiotikaförbrukningen i både forskning och media. Resistenta bakterier kan bli ett hot mot vår framtid om vi inte minskar antibiotikaförbrukningen och vidtar åtgärder för att förhindra smittspridning. Om antalet personer som blir smittade av resistenta bakterier kan reduceras minskar även antibiotikaförbrukningen som i sin tur leder till att färre bakterier utvecklar ett resistensmönster för antibiotika. Detta betyder att det är viktigt att studera och effektivisera hanteringen av smittsamma patienter.   För att reducera antalet smittade patienter måste förebyggande åtgärder vidtas och smittkällan måste kartläggas vid ett upptäckt fall av en smitta. Svensk sjukvård arbetar idag aktivt med smitthantering. Detta begrepp omfattar upptäckt, kontroll och spårning av smitta. Uppdragsgivaren till detta examensarbete, Cambio Healthcare Systems, saknar en fullständig bild över hur smitthanteringen egentligen går till på ett sjukhus. Deras målsättning är att utveckla ett IT-system som kan underlätta smitthanteringsprocessen. Denna studie syftar till att kartlägga informationsflödet vid smitthanteringsprocessen och identifiera de inblandade aktörernas ansvarsområden och skyldigheter enligt regelverket. Vidare syftar arbetet till att presentera åtgärdsförslag som kan minska de identifierade riskerna och effektivisera smitthanteringsprocessen av resistenta bakterier.   För att kartlägga smitthanteringsprocessen genomfördes en fallstudie av ett svenskt universitetssjukhus under våren 2013. Aktörer som studerades var det mikrobiologiska laboratoriet, vårdhygien, smittskyddsenheten, smittskyddsinstitutet samt läkare och sjuksköterskor vid två avdelningar på sjukhuset. Datainsamlingen består av intervjuer, observationer och dokument.   Resultatet av fallstudien visade att smitthanteringsprocessen är ett komplext system med ett omfattande informationsflöde. Huvudaktörerna är vårdhygien, sjukvårdspersonalen och det mikrobiologiska laboratoriet. De är viktiga eftersom deras praktiska handlande är avgörande för att smitthanteringen genomförs. Smittskyddsenheten är inblandad till viss del, men tillhör inte huvudaktörerna. Studien visade även att smittskyddsinstitutet inte hade någon framträdande roll i smitthanteringsprocessen av resistenta bakterier på sjukhuset.   Ansvarsfördelningen är till viss del styrd av smittskyddslagen och enligt denna lag har den behandlande läkaren en central roll i processen. I verkligheten är läkarens roll mindre framträdande vid smittspårningen, vanligtvis delegerar läkaren uppgifter till sjuksköterskor eller till vårdhygien. Mycket av kommunikationen mellan aktörerna är muntlig och detta innebär att ett flertal risker kan uppstå. Vid identifieringen av risker för hela processen konstaterades det att de flesta risker kan uppstå på grund av den mänskliga faktorn, ofta i kombination med användandet av ett otillräckligt datasystem. Åtgärdsförslagen för att effektivisera processen fokuserar därför på att minimera de identifierade riskerna med hjälp av framtida IT-system.   Slutsatsen av studien är att det finns ett stort behov av IT-lösningar för att effektivisera smitthanteringsprocessen av resistenta bakterier. Min rekommendation är att Cambio Healthcare Systems AB bör fokusera på att utveckla ett system för att digitalisera arkiveringen av beläggningslistorna och spåra patientflöden tillbaka i tiden i Cosmic då detta är ett starkt önskemål från kunderna. En annan viktig åtgärd är att utveckla smittspecifika checklistor som visas på datorn i samband med att läkaren får ett positivt provsvar. Slutligen rekommenderar jag Cambio Healthcare Systems AB att utveckla ett smittlarm som kan integreras med deras befintliga whiteboardtavla som nyligen lanserades. / Infectious diseases are a major cost item for the Swedish society. The treatment of infected patients has previously been estimated to 5-10 billion SEK annually and preventive actions cost the Swedish society around one billion SEK every year. Therefore, there are strong economic incentives to reduce the number of infected patients in care, particularly cases caused by resistant bacteria. There is an ongoing debate in both media and research about bacterial resistance and antibiotic consumption. Resistant bacteria can be a threat to our future if we do not reduce the consumption of antibiotics and take measure against infection spreading. If it is possible to reduce the number of resistant bacteria infected patients in the future it enables a decline in antibiotic consumption. This in turn leads to a decreased quantity of bacteria that is able to develop a resistance pattern to antibiotic. Thus, it is highly motivated to study and streamline the process of infection control.   Preventive measures must be taken and the source of the infection must be identified in order to reduce the number of infected patients. The Swedish health care sector is currently working actively with infection control. The concept of infection control encloses the detection, the control and the tracing of the infection. The requestor of this master thesis, Cambio Healthcare Systems AB, does not have a complete picture of the process of the infection control. Their goal is to develop an IT system to facilitate the process of infection control. This study aims to map the information flow of the process and to identify the involved actors’ field of responsibility and obligations according to the law. Further, this thesis aims to present action proposals that can reduce the identified risks and streamline the infection control of resistant bacteria.   A case study of a Swedish university hospital was performed in the spring of 2013 in order to map the process of infection control. The investigated actors were the microbiological laboratory, the local health protection unit (Vårdhygien), the unit of infection control at a regional level (Smittskyddsenheten), the Swedish Institute of Infectious Disease Control (Smittskyddsinstitutet) and physicians and nurses at two hospital departments. The data collection consists of interviews, observations and documents.   The result of this case study shows that the process of infection control is a complex system with an extensive flow of information. The main actors are the local health protection unit, the microbiological laboratory and the medical staff. Their practical actions are essential for the process of infection control. The unit of infection control at a regional level is involved to some extent, but does not belong to the main actors. Furthermore, the study showed that the Swedish Institute of Infectious Disease Control does not have a prominent role in the process at the hospital.   The division of responsibilities is to some extent controlled by the law. According to the law, the physician in charge has a central role in the process of infection control. However, the physician’s role in reality is less prominent. Usually, the physician delegates the tasks to the other actors such as nurses or to the local health protection unit. The communication between the actors is mainly oral and this can cause risks. Most of the identified risks occurred due to human error, often in combination with use of an insufficient IT-system. Therefore, the proposed actions to streamline the process focus on minimizing the identified risks with help of future IT solutions.   The conclusion of this study is that there is a strong demand for IT solutions to streamline the process of infection control of resistant bacteria. My recommendation is that Cambio Healthcare Systems AB should focus on developing a system to digitalize the archiving of the occupancy lists, which also enables tracing the flow of patients back in time. This is a request from several health care professionals. Another important proposed action is to develop a checklist that is specific for every infection disease. Simultaneously as the physician receives the positive test results, this checklist will appear on the physician’s screen. Finally, I recommend Cambio Healthcare Systems AB to develop an alarm to infection diseases that can be integrated with their existing whiteboards that were recently introduced to the market.

The extensive medical and agricultural use and misuse of antibiotics during the last 70 years has caused an enrichment of resistant pathogenic bacteria that now severely threatens our capacity to efficiently treat bacterial infections. While is has been known for a long time that high concentrations of antibiotics can select for resistant mutants, less is known about the lower limit at which antibiotics can be selective and enrich for resistant bacteria. In this thesis we investigated the role of low concentrations of antibiotics and heavy metals in the enrichment and evolution of antibiotic resistance. Selection was studied using Escherichia coli and Salmonella enterica serovar Typhimurium LT2 with different resistance mutations, different chromosomal resistance genes as well as large conjugative multidrug resistance plasmids. Using very sensitive competition experiments, we showed that antibiotic and heavy metal levels more than several hundred-fold below the minimal inhibitory concentration of susceptible bacteria can enrich for resistant bacteria. Additionally, we demonstrated that subinhibitory levels of antibiotics can select for de novo resistant mutants, and that these conditions can select for a new spectrum of low-cost resistance mutations. The combinatorial effects of antibiotics and heavy metals can cause an enrichment of a multidrug resistance plasmid, even if the concentration of each compound individually is not high enough to cause selection. These results indicate that environments contaminated with low levels of antibiotics and heavy metals such as, for example, sewage water or soil fertilized with sludge or manure, could provide a setting for selection, enrichment and transfer of antibiotic resistance genes. This selection could be a critical step in the transfer of resistance genes from environmental bacteria to human pathogens.

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

No description available.

Cattle are commonly treated with antibiotics that may survive digestion and promote antibiotic resistance when manure or composted manure is used as a soil amendment for crop production. This study was conducted to determine the effects of antibiotic administration and soil amendment practices on microbial diversity and antibiotic resistance of bacteria recovered from the surfaces of lettuce and radishes grown using recommended application rates. Vegetables were planted in field plots amended with raw manure from antibiotic-treated dairy cows, composted-manure from cows with different histories of antibiotic administration, or a chemical fertilizer control (12 plots, n=3). Culture-based methods, 16SrDNA amplicon sequencing, qPCR and shot-gun metagenomics were utilized to profile bacteria and characterize the different gene markers for antibiotic resistance. Culture-based methodologies revealed that lettuce grown in soils amended with BSAs had significantly larger clindamycin resistant populations compared to control conditions. Growth in BSAs was associated with significant changes to the bacterial community composition of radish and lettuce. Total sul1 copies were 160X more abundant on lettuce grown in manure and total tet(W) copies were 30X more abundant on radishes grown in manure. Analysis of shotgun metagenomic data revealed that lettuce grown in manure-amended soils possessed resistance genes for three additional antibiotic classes compared to other treatments. This study demonstrates that raw, antibiotic-exposed manure may alter microbiota and the antibiotic resistance genes present on vegetables. Proper composting of BSAs as recommended by the U.S. Department of Agriculture and Environmental Protection Agency is recommended to mitigate the spread of resistance to vegetable surfaces. / MSLFS / Antibiotics are drugs responsible for killing infectious diseases in both humans and animals. In cows, antibiotics are frequently used when they get infections in their udders. These drugs can be excreted through manure and urine and end up in the environment. Manure or composted manure is often applied as a soil amendment for crop production. The presence of antibiotics in soil may promote antibiotic resistance, meaning bacteria that carry antibiotic resistance genes (ARGs) are capable of surviving exposure to drugs that would normally kill them. Such bacteria may eventually pass their ARGs to pathogens, which then could no longer be treated effectively by antibiotics when there is an infection. Thus, there is concern that overuse of antibiotics in agriculture can contribute to reduced effectiveness of antibiotics and the growing global antibiotic resistance health crisis. This study sought to determine if prior antibiotic administration affected the antibiotic resistance of bacteria found on the surfaces of vegetables grown in soil amended with manure or compost from dairy cows. Lettuce and radishes were grown in the field in plots amended with raw manure from antibiotic-treated dairy cows, compost from cows with different histories of antibiotic administration, or a chemical fertilizer control. Mature vegetables were harvested and used to enumerate antibiotic-resistant bacterial colonies. Additionally, the 16S rRNA gene, which is a ubiquitous gene found in all bacteria, was sequenced to identify the kinds of microbes that colonized the radish and lettuce surfaces when grown under the different conditions. DNA was extracted from the bacteria collected from the vegetable surfaces to and different methods were used to identify the kinds of ARGs present and to which kinds of antibiotics they encode resistance. The results of the study indicated that raw, antibiotic-exposed manure may increase the bacteria found on vegetables in addition to their ARGs. Proper composting of manure, as recommended by the U.S. Department of Agriculture (USDA) and the Environmental Protection Agency (EPA), is recommended to mitigate resistance and control microbial populations on fresh vegetables.

Outputs of human-derived chemicals and constituents to the environment, and shifts in these outputs, can result in unintended consequences to human and ecological health. One such shift is the advent of the modern antibiotic era, in which mass production and outputs of antibiotics, which are mostly naturally-derived microbial defense compounds and include a few synthetic antimicrobials, has profound implications for contributing to the spread of antibiotic resistance. Antibiotic resistance arises from mutations and/or sharing of antibiotic resistance genes (ARGs) among bacteria via horizontal gene transfer, with carriage of ARGs by pathogenic bacteria of particular concern to human health. While most attention to stopping the spread of antibiotic resistance has been devoted to the clinic, it is critical to consider the environmental origin, ecology and pathways by which antibiotic resistance spreads in order to develop comprehensive strategies to combat antibiotic resistance. In particular, wastewater treatment plants (WWTPs) represent a potentially key critical control point given that they receive antibiotic resistant bacteria (ARB) and ARGs from the population, which are then routed to activated sludge biological treatment, consisting of high density, highly active microbial populations. The research projects described in this dissertation aimed to explore the occurrence of ARGs in WWTPs, particularly WWTPs in developing countries representing the extremes of what is expected to be encountered in terms of potential to spread antibiotic resistance, and to improve and apply novel technologies for monitoring key markers of antibiotic resistance in WWTPs and affected environments. The pathogen Staphylococcus aureus and a corresponding ARG (methicillin resistance mecA gene) were chosen as model biocontaminants of concern due to their environmental and public health relevance. The results reported in Chapters 3-5 advance the knowledge of bio(nano)sensing techniques and highlight areas of promise and challenge. The results reported in Chapter 2 provided insight into the baseline levels of ARGs expected in a highly impacted WWTP in India, thereby highlighting the magnitude and global scale of the problem of antibiotic resistance as well as the need for innovative solutions. / Ph. D. / Release of human-derived pollutants into the environment can result in unintended consequences to human and environmental health. The rise of antibiotic resistance in disease-causing bacteria serves as a notorious example. Antibiotic resistance arises from mutations and/or sharing of antibiotic resistance genes (ARGs), which are the genetic elements that enable the resistance to occur. While most attention to stopping the spread of antibiotic resistance has been devoted to the clinic, it is critical to consider the environmental factors by which antibiotic resistance spreads in order to develop wellinformed strategies to combat it. In particular, wastewater treatment plants (WWTPs) represent a potentially key critical control point given that they receive antibiotic resistant bacteria (ARB) and ARGs from the population, which are then routed to a highly active biological treatment process. The research projects described in this dissertation aimed to explore the occurrence of ARGs in WWTPs, particularly WWTPs in developing countries representing the extremes of what is expected to be encountered in terms of potential to spread antibiotic resistance, and to improve and apply novel technologies for monitoring key markers of antibiotic resistance in WWTPs and affected environments. The disease-causing bacterium <i>Staphylococcus aureus</i> and a corresponding ARG (methicillin antibiotic resistance mecA gene) were chosen as model biological contaminants of concern due to their environmental and public health relevance. The results reported in Chapters 3-5 advance the knowledge of integrated microbiology and nanotechnology techniques, and also highlight some associated limitations. The results reported in Chapter 2 provide insight into the baseline levels of ARGs expected in a highly impacted WWTP in India, thereby highlighting the magnitude and global scale of the problem of antibiotic resistance as well as the need for innovative solutions.