by Ming-chiu Fung. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1980. / Includes bibliographies.

Chack-yung Yu. / Bibliography: leaves 160-175 / Thesis (M.Phil.) -- Chinese University of Hong Kong, 1983

A key factor in studying evolutionary biology is an understanding of the mechanisms organisms utilise in the ongoing process of adaptation. When faced with a heterogeneous and unpredictable environment, we expect organisms to evolve either as specialists or generalists, yet a unifying theory as to which will evolve is still lacking due to conflicting hypotheses based on limited empirical evidence. Phenotypic plasticity allows a single genotype to express different phenotypes, and has been found as an adaptive response to changing environments in all major taxa. With the advent of genomics it has become possible to study the underlying genetics of this phenomenon. It is however becoming clear that there is no single principle governing the plastic response, but rather a complex set of interactions between what appears to be regulatory and structural genes. With empirical data only recently becoming more readily available, the modelling of plastic responses are often still founded on the theoretical predictions and assumptions for which there is little proof. To bridge the gap between theory and nature, the challenge facing scientists today is the construction of experimental systems where theoretical predictions can be scrutinised. Given that phenotypic plasticity is a widespread phenomenon, understanding the magnitude and constraints of this response is an important issue in the study of evolution. Models have predicted a correlation between genome size and phenotypic plasticity, with increased genome size (complexity) linked to higher levels of phenotypic plasticity. Experimental findings, however, increasingly point to plasticity being governed by complicated sets of interactions between various parts of the genome, the adaptive landscape, and environmental cues. In the work presented here, a study was designed to test for a correlation between genome size and the level of plasticity by, looking at the fitness response of phages exposed to varying temperature. Seven phages differing in genome size and genome composition were used. Genome sizes ranged from 5386 bp to 170 000 bp. Taking advantage of the short generation times of phages, fitness could be measured as the growth rate per hour, which was compared among the different phage groups. The growth of large populations within a constant, controlled environment minimized the complications of environmental heterogeneity, and allowed for quantitative measure of the response to different temperatures. This was used to gain insight into how genome size relates to the level of phenotypic plasticity. Limited generation numbers were allowed for, to ensure population growth could be directly related to the plasticity of the genome, since numerous generations would be required for the effects of selection to become apparent. Adsorption rates are influenced by temperature, and were therefore measured to determine if it had a significant effect on the resulting population density. Results showed a marginal interaction between genome size and phenotypic plasticity, with adsorption rate having no significant effect. More experimental work would be required to verify this finding. / Dissertation (MSc (Genetics))--University of Pretoria, 2006. / Genetics / unrestricted

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

The growth of environmentally persistent biofilms in cooling towers causes several associated problems, including microbiologically-induced corrosion (MIC) and biofouling. Current chemical control methods are not only ineffective against biofilms and costly to procure, they also have downstream environmental impacts when released untreated, or incur additional treatment costs. Bacteriophages are alternative biofilm control agents that have the potential to be more effective, cheaper to produce and yet have a more benign effect on the environment. In this study, biofilms grown under conditions simulating seawater fed cooling towers were characterized and the differences in growth and community make-up across time and different substrates were assessed. An MIC associated bacterium common in cooling tower water, P. aeruginosa, was chosen. Seven bacteriophage strains found to be effective against the chosen bacterium were isolated from wastewater influent. The relative effectiveness of these strains was measured against P. aeruginosa across different salinities. Separate biofilms fed with P. aeruginosa enriched seawater were characterized and the effectiveness of the isolated strains, singly and in cocktails, against the enriched biofilms was measured.

Abstract The use of antibiotics in aquaculture may cause development of antibiotic resistance among pathogens infecting cultured animals and humans. Therefore, the phages were isolated from the culture environment that can infect the pathogen and resistant bacteria. In this study, there were vibriophage and antibacterial phage isolated from CLOZ andSCKF. The small and circle plaque of vibriophage could become striking by decrease in top agar percentage. The electron micrographs of vibriophage and antibacterial phage belonged to the Podoviridae and Myroviridae family. The phages genome could be cut by HidinIII. The different size fragments were compared and matched to similar genome size of phages from NCBI. For the result, vibriophage may belong to the Picovirinae in Podoviridae. The antibacterial phage would be classified into either Mu-like viruses or unclassified Myoviridae. In the infecting test with (103 PFU/ml), the vibriophage lysing the host cell was not evident. Then, infecting with ϕA, ϕ20 and ϕ70 107 PFU/ml), the ϕA could lyse the cell and test the lowest OD after two hours by infecting. ϕ20 lysed the cell at exponential phase and antibacterial phage ϕ70 could lysed the host cell at different ages after six hours by infecting. A could lyse the cell and test the lowest OD after two hours by infecting. ϕ20 lysed the cell at exponential phase and antibacterial phage ϕ70 could lysed the host cell at different ages after six hours by infecting.

Q-Beta phage RNAs with inactivating insertion (8 base) or deletion (17 base) mutations within their replicase genes were transfected into Escherichia coli spheroplasts containing QB replicase provided in trans by a resident plasmid. Replicase-defective (Rep~) Q3 phage produced by these spheroplasts were unable to form plaques on cells lacking this plasmid. When individual Rep~ phage were isolated and grown to high titer in cells containing plasmid derived Q3 replicase, revertant Q3 phage (Rep'), with the original mutation (insertion or deletion) repaired, were obtained at a frequency of ca. 1 x 108. RNA recombination via a "template switching" mechanism involving Q3 replicase, the mutant phage genome, and the plasmid-derived replicase mRNA was shown to be the primary means by which these mutant phages reverted to wild type.

No description available.

Staphylococcal scalded skin syndrome (SSSS) is a life-threatening skin condition caused by Staphylococcus aureus (SA) strains producing exfoliative toxin. Most SSSS cases are associated with Exfoliative Toxin A, encoded by the eta gene and carried by a temperate bacteriophage phiETA (ΦETA). An estimated ~10% of strains containing ΦETA are methicillin-resistant SA (MRSA), raising questions about the factors associated with ΦETA susceptibility. Our study investigates the lysogenization susceptibility of ΦETA in a set of SA strains through lysogenic challenges. To this end, we first isolate phage lysates from a known epidemic strain containing eta. In lysogenic challenge assays, we demonstrate that MRSA strains are resistant to ΦETA acquisition. All the 4/89 (4.5%) strains that were successfully lysogenized with ΦETA were characterized as methicillin-susceptible SA (MSSA). While lysogenic conversion did not modify other phenotypic markers, it increased the biofilm production, indicating potential fitness advantages conferred by ΦETA lysogenization. We also hypothesized that phage sequences in challenged strains may provide immunity against ΦETA, thus prophage presence and diversity may correlate with ΦETA susceptibility. Among the 89 strains tested, we bioinformatically identified that 54 of them contained at least one intact prophage sequence. Interestingly, all successfully lysogenized strains lacked intact prophages in their native state which contradicts the poly-lysogenic nature of SA. Additionally, the enhancement of biofilm formation after lysogenic conversion by ΦETA could be associated with pathogenesis of SSSS, increased invasiveness and even emergence of clinically relevant strains. Further studies are needed to explain the genetic and transcriptional basis of phenotypic changes. Together, our results underscore the vital role of ΦETA prophages in shaping SA diversity and clinical outcomes. This study highlights the necessity of investigating the molecular mechanisms associated with bacteriophage lysogenization and host range expansion.

Les êtres vivants sont confrontés à des parasites qui diminuent leur fitness et se répandent dans la population. En réponse, les hôtes ont développé de nombreuses défenses immunitaires qui sont souvent mises en défaut par l'évolution des parasites. Ces défenses sont de plus souvent extrêmement diversifiées génétiquement. Quel est donc l'apport de la diversité génétique des défenses contre l'évolution des parasites ? Répondre à cette question expérimentalement nécessite un système biologique pour lequel on peut étudier la diversité génétique de l'hôte et l'évolution et la propagation du parasite. Les systèmes bactéries/phages sont de bons candidats pour une telle étude : leur manipulation au laboratoire est aisée, leurs cycles de vie sont rapides et ils ont de forts taux de mutation. La découverte récente de l'immunité CRISPR--Cas a ouvert de nombreuses possibilités : cette dernière a la propriété unique de générer dans le même fond génétique que l'hôte sensible de nombreux allèles de résistance. De plus, son mécanisme de fonctionnement reposant sur une interférence à ARN, la cible d'une résistance est très précisément connue ainsi que les possibilités de la contourner. Ce système permet donc l'étude expérimentale de l'impact de la diversité génétique sur la propagation et l'évolution des parasites, et sur la co-évolution antagoniste. Dans cette thèse, nous cherchons à 1) déterminer l'impact de la composition de la population d'hôtes sur la probabilité qu'une épidémie créée par un virus mutant ait lieu (émergence évolutive), 2) expliciter les causes de l'hétérogénéité de durabilité des résistances et 3) étudier la dynamique co-évolutive entre population génétiquement diversifiée d'hôtes et de parasites. Nous montrons que la composition de la population d'hôtes module fortement la probabilité d'émergence évolutive : une faible diversité génétique associé à un taux intermédiaire d'hôtes sensibles maximisant la probabilité d'émergence évolutive. Dans un second temps, nous montrons que l'immunité CRISPR génère des résistances dont la durabilité est hétérogène et cette hétérogénéité ne peut pas être expliquée par une hétérogénéité des fitness des mutants contournant CRISPR. Enfin, nous montrons que la diversité des résistances est maintenue à court terme par l'hétérogénéité des populations de parasites et que la dynamique co-évolutive est accélérée en présence d'une population génétiquement diverse. Enfin, nous proposons des pistes de recherche qu'il nous parait intéressant d'étudier dans le futur. / Living organisms face parasites which decrease their fitness and spread into their population. In response, hosts have evolved countless immune defenses that are often circumvented by parasite evolution. These defenses are usually extremely diverse. What is the impact of such genetic diversity on the protection against the evolution of parasites? Answering this question experimentally requires an experimental system in which host genetic diversity and parasite evolution and spreading can be monitored. Phages and bacteria systems are ideal candidates for such studies as their handling is easy in the lab, their life cycle is short and their mutation rates is high. The recent discovery of CRISPR--Cas immunity has opened many possibilities. Indeed, this immunity has the unique property to generate in the same genetic background as the sensitive host, numerous resistant alleles. In addition, it relies on an interference--RNA-like pathway, which results in the precise understanding of phage bypassing and in the ability to predict the targeted sequence. This system hence allows the experimental study of the impact of host genetic diversity on the epidemiology and the evolution of parasites and on antagonist coevolution. In this PhD, we 1) study how the host population composition impacts the probability of an epidemic created by an escape mutant (evolutionary emergence), 2) try to understand the causes of the heterogeneity in durability of resistances and 3) monitor the coevolution dynamic between genetically diverse populations. We show that the composition of the host population impacts the probability of evolutionary emergence: a low resistances diversity with an intermediate proportion of sensitive hosts maximises the probability of evolutionary emergence. Second, we show that CRISPR--Cas resistances are heterogeneous in their durability and this is not explained by the heterogeneity of escape mutants fitness. Third, we show that resistances diversity is conserved in a short term by parasites genetic diversity and that the coevolutionary dynamic is fastened by parasite intra-specific genetic diversity. Finally, we discuss research questions that we find interesting to develop in the near future.