This study is entitled, “determination of the presence of antibiotics and bacterial pathogens with their susceptibility in fresh milk from the San Nicolas community and the cities of Penipe, Guano, and Chambo in the Chimborazo province”. It was funded by the Benson Institute of Brigham Young University and Nestle. The objectives were: 1) to determine the presence of antibiotics in fresh milk from the San Nicolas community and the cities of Penipe, Guano, and Chambo, 2) to determine the presence of bacterial pathogens in milk from the places previously mentioned, and 3) to detect the susceptibility of the isolated bacteria to the most commonly used antibiotics on the market. With regard to the presence of antibiotics, they were only found once in Chambo (beta-lactams) and once in San Nicolas (tetracycline). Three types of pathogenic bacteria were found: Escherichia coli, Streptococcus agalactiae, and Staphylococcus aureus. Only one bacterium was found that has not been studied: Pseudomonas aureoqinosa. This bacterium was found in milk from San Nicolas. With regard to the susceptibility of the bacteria to antibiotics, it was found that Escherichia coli was sensitive to the following antibiotics: nitrofurans, erimethoprina, trimethoprim and sulfonamides, and tetracyclines and spectomicina. Escherichia coli also showed medium sensitivity to chloramphenicol as well as resistance to ampicillin and sulfonamides. Streptococcus agalactiae was sensitive to lincomycin, neomycin, enrofloxacin, and clindamycin. It also showed medium sensitivity to erythromycin and low tetracycline and penicillin as well as resistance to gentamicin. Lastly, Staphylococcus aureus was sensitive to neomycin and enrofloxacin. In addition, it showed medium sensitivity to lincomycin, gentamicin, clindamycin, and tetracycline as well as resistance to low erythromycin and penicillin. This study was done over a four month period with at least 12 samples being collected from each area. A total of 60 samples were analyzed. Based on the results obtained, our recommendation is that hygienic measures are taken in obtaining and transporting milk to eliminate the growth of bacterial pathogens. Also, we recommend the use of the effective antibiotics listed above in order to completely eradicate these microorganisms.

Vibrio salmonicida causes cold-water vibriosis in salmon populations around the world and causes financial damage to fisheries designed to farm these salmon. Very little is known about the physiology of how V. salmonicida causes disease and measures to contain vibriosis are restricted to either vaccinating individual fish against disease or administering antibiotics when an outbreak is detected. These procedures are costly and increase the risk for selection of antibiotic-resistant V. salmonicida strains. A recent reoccurrence of outbreaks in Norwegian fisheries provided incentive to better understand the virulence mechanisms of V. salmonicida. In this thesis, a proteomic approach was used to identify proteins that were differentially expressed when cells were grown in vitro under simulated virulence conditions (i.e. 5˚C and in the presence of exogenously supplied autoinducer 3-oxo-hexanoyl-homoserine lactone). Some examples of proteins with significantly altered expression that stood out at as homologs of potential virulence factors were: an exported serine protease DegQ, a multi-drug transporter HlyD, and an outer membrane protein OmpU. The proteomic approach allowed us to identify large numbers of proteins that are expressed by V. salmonicida, facilitating hypothesis-driven research in order to support possible roles for some of these proteins in virulence

The main challenge in studying genetics has evolved from identifying variations and their impact on traits to comprehending the molecular mechanisms through which genetic variations affect human biology, including disease susceptibility. Despite having identified a vast number of variants associated with human traits through large scale genome wide association studies (GWAS) a significant portion of them still lack detailed insights into their underlying mechanisms [1]. Addressing this uncertainty requires the development of precise and scalable approaches to discover how genetic variation precisely influences phenotypes at a molecular level. In this study, we developed a pipeline to automate the annotation of structural variant feature effects. We applied this pipeline to a dataset of 33,942 variants from the ClinVar and GnomAD databases, which included both pathogenic and benign associations. To bridge the gap between genetic variation data and molecular phenotypes, I implemented Non-negative Matrix Factorization (NMF) on this large-scale dataset. This algorithm revealed 6 distinct clusters of variants with similar feature profiles. Among these groups, two exhibited a predominant presence of benign variants (accounting for 70% and 85% of the clusters), while one showed an almost equal distribution of pathogenic and benign variants. The remaining three groups were predominantly composed of pathogenic variants, comprising 68%, 83%, and 77% of the respective clusters. These findings revealed valuable insights into the underlying mechanisms contributing to pathogenicity. Further analysis of this dataset and the exploration of disease-related genes can enhance the accuracy of genetic diagnosis and therapeutic development through the direct inference of variants that are likely to affect the functioning of essential genes.

Trypanosoma brucei and related parasites are causative agents of severe diseases that affect global health and economy. T. brucei is responsible for sleeping sickness in humans (African trypanosomiasis) and a wasting disease in livestock. More than 100 years after T. brucei was identified as the etiological agent for sleeping sickness, available treatments remain inadequate, complicated by toxicity, lengthy and expensive administration regiments, and drug-resistance. There is clear need for the development of a new antitrypanosomal drugs. Due to the unique evolutionary position of these early diverging eukaryotes, trypanosomes posses a number of biological properties unparalleled in other organisms, including humans, which could prove valuable for new drug targets. One of the most distinctive properties of trypanosomes is their mitochondrial DNA, called kinetoplast DNA (kDNA). kDNA is composed of over five thousand circular DNA molecules (minicircles and maxicircles) catenated into a topologically complex network. Replication of kDNA requires an elaborate topoisomerase-mediated release and reattachment mechanism for minicircle theta structure replication and at least five DNA polymerases. Three of these (POLIB, POLIC, and POLID) are related to bacterial DNA polymerase I and are required for kDNA maintenance and growth. Each polymerase appears to make a specialized contribution to kDNA replication. The research described in this dissertation is a significant contribution to the field of kDNA replication and the advancement of kDNA replication proteins as putative drug targets for sleeping sickness. Functional characterization of POLIB indicated that it participates in minicircle replication but is likely not the only polymerase contributing to this process. Gene silencing of POLIB partially blocked minicircle replication and led to the production of a previously unidentified free minicircle species, fraction U. Characterization of fraction U confirmed its identity as a population of dimeric minicircles with non-uniform linking numbers. Fraction U was not produced in response to silencing numerous other previously studied kDNA replication proteins but, as we demonstrated here, is also produced in response to POLID silencing. This common phenotype led us to hypothesize that POLIB and POLID both participate in minicircle replication. Simultaneously silencing both polymerases completely blocked minicircle replication, supporting a model of minicircle replication requiring both POLIB and POLID. Finally, we demonstrate that disease-causing trypanosomes require kDNA and the kDNA replication proteins POLIB, POLIC, and POLID. These data provide novel insights into the fascinating mechanism of kDNA replication and support the pursuit of kDNA replication proteins as novel drug targets for combating African trypanosomiasis.

京都大学 / 新制・課程博士 / 博士(工学) / 甲第24898号 / 工博第5178号 / 新制||工||1988(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 伊藤 禎彦, 教授 松田 知成, 教授 越後 信哉 / 学位規則第4条第1項該当 / Doctor of Philosophy (Engineering) / Kyoto University / DGAM

Plant roots are associated with a diverse microbial community of bacteria and fungi. The root microbiome communities associate with the root surface or penetrate the superficial layers of root tissues. It is of high agronomic interest to characterize root microbiomes and identify microbes with beneficial effects on plant defense against pathogens. Soybean taproot decline (TRD) is an emerging soil-borne disease caused by a fungus classified as Xylaria necrophora, challenging to control and with devastating effects on yield. Our group has initiated a study of the soybean root microbiome with three primary experiments: (I) perform a comparative study of root microbiomes and mycobiomes from healthy and TRD-affected soybean, (II) isolate, screen and (III) identify soybean root bacterial endophytes with anti-X. necrophora and anti-TRD properties. Our long-term objective is to discover and implement microbiome-based management techniques that enhance the health and yield of soybeans.

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Avian pathogenic Escherichia. coli (APEC) and Campylobacter are pathogenic threats to poultry and human health, respectively. In this study, the prevalence of these pathogens in Mississippi broilers and their antimicrobial resistance (AMR) properties were investigated, and a multidrug-resistant APEC strain (APEC-O2-MS1170) was further explored by whole-genome sequencing (WGS). The efficacy of in ovo injection of Lactobacillus in reducing the APEC in broilers was evaluated. Results revealed a high prevalence of APEC and Campylobacter in broilers and broiler products. A lot of isolates were resistant to antibiotics of different sorts. Moreover, the in ovo administration of Lactobacillus did not reduce the incidence of APEC. The WGS of APEC-O2-MS1170 revealed its detailed AMR and virulence properties and alerted a potential zoonotic risk. In conclusion, the Lactobacillus did not reduce the incidence of APEC in broilers, and the prevalence and AMR of APEC and Campylobacter are still challenges faced by the poultry industry.

No description available.