Antibiotic resistance is a global threat that can severely imperil public health. To curb the spread of antibiotic resistance, it is imperative that efforts commensurate with a “One Health” approach are undertaken. Given that interconnectivities among ecosystems can serve as conduits for the proliferation and dissemination of antibiotic resistance, it is increasingly being recognized that a robust global environmental surveillance framework is required to promote One Health. The ideal aim would be to develop approaches that inform global distribution of antibiotic resistance, help prioritize monitoring targets, present robust data analysis frameworks to profile resistance, and ultimately help build strategies to curb the dissemination of antibiotic resistance. The work described in this dissertation was aimed at evaluating and developing different data analysis paradigms and their applications in investigating and characterizing antibiotic resistance across different resistomes. The applications presented in Chapter 2 illustrate challenges associated with various environmental data types (especially metagenomics data) and present a path to advance incorporation of data analytics approaches in Environmental Science and Engineering research and applications. Chapter 3 presents a novel approach, ExtrARG, that identifies discriminatory ARGs among resistomes based on factors of interest. The results in Chapter 4 provide insight into the global distribution of ARGs across human fecal and sewage resistomes across different socioeconomics. Chapter 5 demonstrates a data analysis paradigm using machine learning algorithms that helps bridge the gap between information obtained via culturing and metagenomic sequencing. Lastly, the results of Chapter 6 illustrates the contribution of phages to antibiotic resistance. Overall, the findings provide guidance and approaches for profiling antibiotic resistance using metagenomics and machine learning. The results reported further expand the knowledge on the distribution of antibiotic resistance across different resistomes. / Antibiotic resistance is a global threat that can severely imperil public health. To curb the spread of antibiotic resistance, it is imperative that efforts commensurate with a "One Health" approach are undertaken. Given that interconnectivities among ecosystems can serve as conduits for the proliferation and dissemination of antibiotic resistance, it is increasingly being recognized that a robust global environmental surveillance framework is required to promote One Health. The ideal aim would be to develop approaches that inform global distribution of antibiotic resistance, help prioritize monitoring targets, present robust data analysis frameworks to profile resistance, and ultimately help build strategies to curb the dissemination of antibiotic resistance. The work described in this dissertation was aimed at evaluating and developing different data analysis paradigms and their applications in investigating and characterizing antibiotic resistance across different resistomes. The applications presented in Chapter 2 illustrate challenges associated with various environmental data types (especially metagenomics data) and present a path to advance incorporation of data analytics approaches in Environmental Science and Engineering research and applications. Chapter 3 presents a novel approach, ExtrARG, that identifies discriminatory ARGs among resistomes based on factors of interest. The results in Chapter 4 provide insight into the global distribution of ARGs across human fecal and sewage resistomes across different socioeconomics. Chapter 5 demonstrates a data analysis paradigm using machine learning algorithms that helps bridge the gap between information obtained via culturing and metagenomic sequencing. Lastly, the results of Chapter 6 illustrates the contribution of phages to antibiotic resistance. Overall, the findings provide guidance and approaches for profiling antibiotic resistance using metagenomics and machine learning. The results reported further expand the knowledge on the distribution of antibiotic resistance across different resistomes. / Doctor of Philosophy / Antibiotic resistance is ability of bacteria to withstand an antibiotic to which they were once sensitive. Antibiotic resistance is a global threat that can pose a serious threat to public health. In order to curb the spread of antibiotic resistance, it is imperative that efforts commensurate with the "One Health" approach. Since ecosystem networks can act as channels for the spread and spread of antibiotic resistance, there is growing recognition that a robust global environmental monitoring framework is required to promote a true one-health approach. The ideal goal would be to develop approaches that can inform the global spread of antibiotic resistance, help prioritize monitoring objectives and present robust data analysis frameworks for resistance profiling, and ultimately help develop strategies to contain the spread of antibiotic resistance. The objective of the work described in this thesis was to evaluate and develop different data analysis paradigms and their applications in the study and characterization of antibiotic resistance in different resistomes. The applications presented in Chapter 2 illustrate challenges associated with various environmental data types (especially metagenomics data) and present a path to advance incorporation of data analytics approaches in Environmental Science and Engineering research and applications. The Chapter 3 presents a novel approach, ExtrARG, that identifies discriminatory ARGs among resistomes based on factors of interest. The chapter 5 demonstrates a data analysis paradigm using machine learning algorithms that helps bridge the gap between information obtained via culturing and metagenomic sequencing. The results of Chapters 4 provide insight into the global distribution of ARGs across human fecal and sewage resistomes across different socioeconomics. Lastly, the results of Chapter 6 illustrates the contribution of phages to antibiotic resistance. Overall, the findings provide guidance and approaches for profiling antibiotic resistance using metagenomics and machine learning. The results reported further expand the knowledge on the distribution of antibiotic resistance across different resistomes.

Les rejets urbains par temps de pluie dégradent l’état écologique des écosystèmes aquatiques et peuvent induire une exposition des populations humaines aux contaminants chimiques et microbiens (bactéries, virus, parasites). L’objectif de ce travail de thèse était d’évaluer les effectifs et de prédire le devenir de bactéries pathogènes introduites dans les milieux aquatiques par une source majeure comme les eaux usées rejetées par des dispositifs tels que les déversoirs d’orage (DO) et les lagunes d’épuration (WWTL). La répartition d’un agent pathogène fortement liée aux milieux hydriques, Pseudomonas aeruginosa, a été comparée avec celles observées pour des indicateurs de contaminations fécales (E. coli et les entérocoques intestinaux), mais également avec celle de l’espèce pathogène Aeromonas caviae. La dangerosité des formes retrouvées dans ces milieux a été évaluée par approches moléculaires (PFGE et MLST). Les résultats obtenus montrent un fort apport en P. aeruginosa via les eaux usées, avec un effet significatif sur les effectifs observés en fonction de l’intensité des pluies et des périodes de temps sec, et les fluctuations du régime hydrologique et des paramètres physico-chimiques. Une grande diversité infra-spécifique des P. aeruginosa, et la capacité de certains génotypes à s’installer durablement dans ces milieux (macrophytes et périphyton) ont été observées. Certaines souches ont par ailleurs montré une parenté avec des lignées d’infections communautaires, ou encore des clones épidémiques majeurs (PA14 et C). Des études en microcosme ont été effectuées pour valider les interactions observées avec certains macrophytes, et identifier des propriétés d’adhérence bactérienne (dont les lectines) impliquées dans ces interactions. Ces travaux ont impliqué une analyse de la distribution des gènes lecA et lecB, codant des lectines chez P. aeruginosa, et une étude de leurs ligands. Le gène lecA a été localisé dans une zone de forte plasticité génomique. Ces travaux ont permis la description d’une nouvelle structure de l’adhésine LecB / Urban wet-weather discharges degrade the ecological status of aquatic ecosystems and may expose human populations to chemical and microbial contaminants (bacteria, viruses, parasites). The aim of this thesis was to evaluate the numbers and predict the fate of pathogenic bacteria introduced into aquatic ecosystems by a major source like wastewater from devices such as combined sewer overflows (CSO) and wastewater treatment lagoons (WWTL). The distribution of a human pathogen closely linked to hydric environments, Pseudomonas aeruginosa, was compared with those observed for fecal indicators (E. coli and intestinal enterococci), but also with that of Aeromonas caviae pathogenic species. Dangerousness of strains found in these environments was evaluated by molecular approaches (PFGE and MLST). Obtained results showed a high contribution of wastewater in P. aeruginosa release, with a significant effect of rainfall intensity and preceding dry periods, in addition to changes in hydrological regime and physico-chemical parameters on recorded data. A large infra-specific diversity was observed within P. aeruginosa and the ability of some genotypes to colonize permanently aquatic surfaces (macrophytes and periphyton) were observed. Some strains showed a kinship with lineages of community infections or major epidemic clones (PA14 and C). Microcosm studies were performed to validate observed interactions with macrophytes, and to identify bacterialadhesion properties (including lectins) involved in these interactions. These investigations involved analysis of the distribution of lectin encoding loci lecA and lecB within P. aeruginosa, and a study of their ligands. lecA was located in a highly unstable genomic region. This work allowed the description of a new structure of the adhesin LecB

En santé publique, de nouveaux programmes de surveillance et de gestion sont à proposer notamment pour les masses d'eaux fortement affectées par l'urbanisation et les rejets urbains par temps de pluie. Les niveaux, les réservoirs et les sources de contamination microbiologique des milieux aquatiques doivent être évalués et identifiés, et leur incidence sur la dissémination des bactéries pathogènes et les risques d'exposition des populations humaines comprises. Parmi les agents pathogènes retrouvés dans les milieux hydriques, Pseudomonas aeruginosa représentent une préoccupation sanitaire majeure. A partir de deux sites expérimentaux, les objectifs de ce travail de thèse furent de définir la contribution des rejets d'eaux usées sur la prévalence de P. aeruginosa dans les cours d'eau récepteurs et d'étudier l'écologie des formes introduites dont ces habitats incluant leur dynamique spatiotemporelle. Les sédiments, les biofilms de surface (périphyton) et les végétaux aquatiques submergés, permettraient leur survie voire la multiplication de certains génotypes de Pseudomonas aeruginosa. La répartition des indicateurs de contaminations fécales et de la bactérie pathogène Aeromonas caviae ont également été étudiée. Il a été mis en évidence que les forces hydrauliques, le morpho-dynamisme de la rivière et les variations saisonnières étaient des facteurs structurants de la répartition des contaminants microbiens analysés. La dangerosité des souches isolées a été évaluée et montré que toutes les souches avaient un potentiel de virulence élevée. Ceci s'est confirmé par la détection de clones épidémiques majeurs au sein de la collection dont des souches apparentées aux clones PA14 ou C. Les capacités métaboliques de ces souches ont été étudiées dont leur antibio-résistance / In Public Health, new monitoring and management programs are needed, especially for the aquatic environments strongly affected by urbanization and urban wet weather discharges. The levels, reservoirs and sources of microbiological contamination of the aquatic environment should be assessed and identified, and their impact on the spread of pathogenic bacteria and the risk of exposure of human populations understood. Among the pathogens found in water environments, Pseudomonas aeruginosa is of major health concern. From two experimental sites, the objectives of this work were to define the contribution of wastewater discharges on the prevalence of P. aeruginosa in receiving watercourses and study the ecology of the introduced forms, including their spatiotemporal dynamic and preferential habitats. Sediments, surface biofilms (periphyton) and the submerged aquatic vegetations appeared to favour the survival or growth of some genotypes of Pseudomonas aeruginosa. The distribution of fecal indicator bacteria and of Aeromonas caviae were also studied. It was highlighted that hydraulic forces, the morpho-dynamics of the river and the seasonal vaiations were determinant factors in the distribution of the analyzed microbial contaminants. The health hazard of the clones found in these systems was estimated through indirect molecular approaches. It was shown that all Pseudomonas aeruginosa strains had a high virulence potential and that some were related to the PA14 and C clones which are spread worldwide and pathogenic

No controle da qualidade da água, parâmetros microbiológicos são avaliados e devem estar de acordo com os limites estipulados em portarias e resoluções. Todas as metodologias de monitoramento microbiano demandam o cultivo dos organismos-alvo. Em muitos casos, as cepas isoladas devem ser analisadas por teses fenotípicos para determinação da espécie. Por meio de inóculos de E. coli, demonstrou-se que a variação na técnica de plaqueamento se deve especialmente à falta de consistência entre as diluições. Além disso, para a análise de réplicas, comarando-se as médias de dois métodos de diluição, foi demonstrado que ser utilizado apenas uma série de diluição derivada de um único inóculo. Utilizando-se culturas puras de E. coli, E. faecalis e P. aeruginosa, a recuperação foi similar entre os meios seletivos respectivos (m-Endo, m-EI, m-PA-C) e meio não seletivo (Tripticase Soy Agar). A utilização da técnica de membrana filtrante resultou em contagens significativamente maiores para E.coli e E. faecalis em comparação ao método de spread plate. A microbiota natural presente na água potável (torneira) não influenciou significativamente as contagens de E. coli, E. faecalis e P. aeruginosa em meios seletivos. Entretanto, na água mineral engarrafada inoculada artificialmente com P. aeruginosa, a contagem foi significativamente maior na réplica estéril. Na análise de água marinha, e na réplica não estéril inoculada com E. faecalis, a contagem foi signifivativamente menor e as células de P. aeruginosa produziram colônias atípicas. Analisando-se amostras do meio ambiente (biofilmes de estação de tratamento de água, lodo de esgoto e água de córrego contaminado), 45 colônias típicas isoladas em meios de cultura seletivos foram identificadas por meio do sequenciamento do gene 16S rRNA, testes bioquímicos e MALDI-TOF. Os resultados de sequenciamento do gene 16SrRNA tiveram baixa correlação com a identificação dos organismos por testes bioquímicos (46,6% em nível de gênero e 20% em nível de espécie) e MALDI-TOF (60% em nível de gênero e 48,8% em nível de espécie). Para as cepas identificadas como E. coli e Enterococcus spp., a correlação entre o sequenciamento e MALDI-TOF foi de 75% e 62%, respectivamente. Uma vez que a quantificação de micro-organismo por membrana filtrante depende do micro-organismo em análise e do tipo de água, é necessário que os laboratórios realizem testes de padronização antes de implementar essa metodologia. Os resultados demonstram que os métodos convencionais utilizados não são adequados e suficientes para a análise da qualidade da água e, portanto, novas metodologias devem ser empregadas. Idealmente devem ser utilizadas técnicas baseadas em características fenotípicas, genômica e proteômica cujos resultados são complementares e fornecem uma identificação mais acurada. / All over the world regulatory agencies specify microbiological water quality parameters to guarantee water safety. Conventional microbiological water quality analysis is based on the cultivation of the target organisms. In many analysis protocols, phenotypic assays of isolated strains are mandated for species determination. Reference samples are required for quality assessment and quality control of analytical protocols. Using E. coli as a model organism, it was demonstrated here that the variability of plate counts of reference cultures is caused mainly by the spread of counts in individual serial dilutions. Besides, comparing the means obtained by two dilution approaches, it was demonstrated that in the analysis of replicates it is possible to use only a set of dilutions derived for a unique inoculum. Recovery of pure cultures of E. coli, E. faecalis and P. aeruginosa was similar on selective culture media (m-Endo, m-EI, m-PA-C) and with the non-selective medium (Tripticase Soy Agar). The membrane filter technique yielded significantly higher counts for E.coli and E. faecalis in comparison to spread plate method. The autochthonous microbiota of potable water (tap water) did not influence the counts of E. coli, E. faecalis and P. aeruginosa on selective culture media. However, the sterile replica of mineral water spiked with P. aeruginosa showed higher counts for these bacteria. For marine water analysis, the non-sterile replica spiked with E. faecalis yielded low counts and P. aeruginosa produced an atypical colony. Both, sample-induced variation in target strain recovery and colony appearance on culture plates indicates the requirement for method evaluation tests on particular sample matrices before implementing routine microbiological analysis by culture-based methods in the laboratory. 45 typical colonies obtained from environmental samples in selective culture media (biofilms from activated sludge, drinking water treatment plants and water from creek contaminated with raw sewage) were analyzed by 16S rRNA gene sequencing, biochemical phenotypic assays and MALDI-TOF. Sequencing showed low correlation with phenotypic assays (46,6% at the genus level and 20% at the species level) and MALDI-TOF (60% at the genus level and 48,8% at the species level). On the other hand, the strains identified as E. coli an Enterococcus spp. by 16S rDNA gene sequencing demonstrated a high correlation with MALDI-TOF (75% and 62%, respectively). Since the results showed that conventional methods aren´t suitable and sufficient to assess water quality, new technologies should be employed. Ideally, techniques should be based on phenotypic, genomic and proteomic features since the results are complementary and provide a more accurate identification.

The savannas of southern Africa are a highly variable and globally-important biome supporting rapidly-expanding human populations, along with one of the greatest concentrations of wildlife on the continent. Savannas occupy a fifth of the earth's land surface, yet despite their ecological and economic significance, understanding of the complex couplings and feedbacks that drive spatiotemporal patterns of change are lacking. In Chapter 1 of my dissertation, I discuss some of the different theoretical frameworks used to understand complex and dynamic changes in savanna structure and composition. In Chapter 2, I evaluate spatial drivers of water quality declines in the Chobe River using spatiotemporal and geostatistical modeling of time series data collected along a transect spanning a mosaic of protected, urban, and developing urban land use. Chapter 3 explores the complex couplings and feedbacks that drive spatiotemporal patterns of land cover (LC) change across the Chobe District, with a particular focus on climate, fire, herbivory, and anthropogenic disturbance. In Chapter 4, I evaluated the utility of Distance sampling methods to: 1) derive seasonal fecal loading estimates in national park and unprotected land; 2) provide a simple, standardized method to estimate riparian fecal loading for use in distributed hydrological water quality models; 3) answer questions about complex drivers and patterns of water quality variability in a semi-arid southern African river system. Together, these findings have important implications to land use planning and water conservation in southern Africa's dryland savanna ecosystems. / Ph. D.

Anthropogenic activities generate waste products that pollute the environment with bacteria and heavy metals. This research assessed pollution of the Kahwa River, Bukavu Town, DRC with cadmium and lead (HMs) and bacterial enteropathogens. A survey of businesses, households and healthcare facilities showed general use of the river to remove effluent and waste. Indicator organisms were cultured at over 200 cfu/100 ml showing faecal contamination of the river water. Antibiotic resistance was shown by enteropathogenic Vibrio cholerae and Salmonella typhi to ampicillin and cotrimoxazole with some sensitivity shown to ciprofloxacin. River water contained HMs at around 40 times the World Health Organisation limit for drinking water. The bacteria, particularly from river sediment, tolerated HMs up to a concentration of 1.5 mg/ml. The presence in the Kahwa River of antibiotic-resistant pathogens showing tolerance to HMs has serious public health implications / Environmental Management / M.Sc. (Environmental management)

Anthropogenic activities generate waste products that pollute the environment with bacteria and heavy metals. This research assessed pollution of the Kahwa River, Bukavu Town, DRC with cadmium and lead (HMs) and bacterial enteropathogens. A survey of businesses, households and healthcare facilities showed general use of the river to remove effluent and waste. Indicator organisms were cultured at over 200 cfu/100 ml showing faecal contamination of the river water. Antibiotic resistance was shown by enteropathogenic Vibrio cholerae and Salmonella typhi to ampicillin and cotrimoxazole with some sensitivity shown to ciprofloxacin. River water contained HMs at around 40 times the World Health Organisation limit for drinking water. The bacteria, particularly from river sediment, tolerated HMs up to a concentration of 1.5 mg/ml. The presence in the Kahwa River of antibiotic-resistant pathogens showing tolerance to HMs has serious public health implications / Environmental Management / M.Sc. (Environmental management)