The most successful antimicrobial agents in clinical use are of microbial origin and of these the greatest variety has been found in the genus Streptomyces. However, the resistance of the pathogenic microbes to the commonly used antibiotics is increasing as a result of the wide-spread and long-term use of these antibiotics. Therefore, understanding the strategies that bacteria use to become resistant is of crucial need. Streptomycetes are Gram positive bacteria, commonly found in soil and are known antibiotic-producers. The focus of this thesis was to underpin the mechanism of resistance to penicillin G in isolated strains of streptomycetes that exhibit elevated resistance to penicillin G and to characterise these organisms. Moreover, to investigate the interaction between penicillin G and PBPs in Streptomyces strains and investigate the relationship between growth rate and penicillin G resistance in Streptomyces in vitro. Ninety six Streptomycetes were isolated and characterized. Morphological examination and the16s rRNA sequences of these strains indicated that strains belong to the species Streptomyces. The MICs and MBCs for penicillin G for the isolated Streptomyces strains were measured by plate culture. Some strains showed growth up to 400 μg/ml with penicillin G, which indicate that the strains were highly resistant against penicillin G. Some strains were unable to grow at penicillin concentrations above 200μg/ml. Also, The MICs of penicillin G for isolated Streptomyces strains were measured using a novel OxoPlates® system in 96-well culture format employing Mueller-Hinton broth culture. The MICs of all strains ranged from 1-100 μg /ml. Results indicate that the sensitivity of Streptomyces strains of penicillin G is not directly related to β-lactamase production in the panel of isolates examined. There was no correlation between the MICs of penicillin G and the growth rate in these isolates. Likewise, there was no association between the position of beta-lactamase producing and non-beta-lactamase producing strains on the phylogenetic tree and their beta-lactamase xii activity. Beta-lactamase producing and non-producing strains refers to the same ancestral origin clade. Additionally, the comparative analysis of 16S rRNA gene sequence and phylogenetic relationship of strain (W43) revealed that the isolate clustered with (W76) Streptomyces lividans strain YLA0. Bocillin (a penicillin binding protein stain) staining in β-lactamase producing strains showed staining throughout the mycelia whereas in non β-lactamase producing strains staining only occurred in certain parts of the mycelia. Bocillin also revealed that in spores PBPs were located on both poles of the spores. Streptomyces coelicolor has the ability to grow at high concentrations of penicillin G up to 640 μg/ml in continuous culture. It also has the capacity to grow at very low amounts of dissolved oxygen in continuous culture. Significantly, there was a correlation between the growth rate of S. coelicolor and the resistant to penicillin G. S. Coelicolor was more sensitive to penicillin G at a high dilution rate. Furthermore, our strategy of using the Bug-Lab for monitoring the progress of S. Coelicolor 1147 in continuous culture, even at low concentrations of cells in real time was successful.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:697527 |
| Date | January 2016 |
| Creators | Alkut, W. |
| Contributors | Hobbs, G. ; Evans, K. ; Foulkes, J. |
| Publisher | Liverpool John Moores University |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://researchonline.ljmu.ac.uk/4613/ |
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