The growing threat of drug- resistant Staphylococcus aureus (S. aureus) infections mandates the need to develop novel, effective and alternative antibacterial therapeutics. Despite infection prevention and control measures, methicillin resistant S. aureus (MRSA)-associated deaths reached 11,285 in 2011 in the USA (CDC, 2013). To counteract the threat of drug resistant S. aureus, we sought to construct and characterize a novel therapeutic based on the display of lytic antibacterial enzymes, termed endolysins. These endolysins were displayed on the surface of a specific bacterial virus, bacteriophage (phage), to generate lytic antibacterial nanoparticles. Endolysins are encoded individually by a variety of double-stranded DNA phage and act to direct host lysis and escape. These lytic enzymes confer a high degree of host specificity that could potentially substitute for, or be combined with, antibiotics in the treatment of gram-positive drug resistant bacterial infections such as MRSA. In this study, modular domains of the phage-encoded endolysin K enzyme, specific to S. aureus, were displayed on the capsid surface of phage lambda () via fusion with the λ major head (capsid) protein, gpD. The constructs of displayed endolysins were prepared in various combinations to maximize the functional display of gpD::X fusions on the phage. Phage lysates were generated, collected and purified and lysis was investigated by adding to fresh lawns of MRSA, vancomycin resistant S. aureus (VRSA) and bovine S. aureus. Phage preparations did not readily confer cell lysis, likely due to poor incorporation of the fusions onto the functional phage capsid. We purified the fusion proteins (gpD::X) and tested them for their lytic activity. We noted that the activity of the gpD::LysK protein was not impaired by the fusion and demonstrated lysis on live and dead (autoclaved) bovine S. aureus. In contrast to gpD::LysK, the gpD::CHAP protein fusion, expressing only the CHAP catalytic domain of endolysin K showed variable results in the lysis assays that we performed. In the zymogram assay, gpD::CHAP did not elicit any observable lysis on live bovine S. aureus cells, but did effectively lyse dead cells of the same S. aureus species; however, it was highly lytic in the inhibition assay on bovine S. aureus. The CHAP::gpD protein fusion, which is the CHAP domain fused to the N terminus of gpD only showed its ability to inhibit bovine S. aureus growth on the inhibition assay. The fusion of endolysin K or its CHAP domain to gpD protein does not seem to interfere with lytic activity, but may result in recalcitrant gpD fusions that compromise the ability to efficiently decorate the phage capsid. Suggestions for improved fusion capsid integration are discussed.

Staphylococcus aureus is the most clinically important Staphylococcus species and is associated with high mortality in patients with positive blood cultures. S. aureus bacteria may cause a variety of disease manifestations ranging from minor skin infections to life-threatening conditions such as pneumonia, meningitis, osteomyelitis, endocarditis, toxic shock syndrome (TSS) and sepsis. This microorganism belonging to the gram positive cocci may also be part of the normal flora. In Sweden, penicillinase-stable penicillins are the primary alternatives to treat S. aureus infection. Mutations in genes encoding the penicillin binding proteins (PBP2) in the bacteria which lead to a lower affinity for the  beta-lactam antibiotics define  methicillin resistant S. aureus (MRSA) which is a significant global health problem. Other resistance mechanisms of S. aureus are present, and one of these is penicillinase production which is associated with resistance to penicillin G. In order to detect penicillinase production in S. aureus, there are several methods but the European guidelines recommend disc diffusion and the clover-leaf test for follow-up if the zone diameter for benzylpenicillin (PcG) is 26 mm or more. There are no modern Swedish studies on the prevalence of S. aureus susceptible to PcG and this has recently attained interest from infectious disease physicans. Thus, the purpose of this study was to investigate the frequency of S. aureus susceptible to PcG from blood cultures isolated during 2012 from the Kalmar county.    Disc diffusion testing showed that 32% of 90 unique isolates tested had an inhibition zone diameter of PcG that was ≥ 26 mm in diameter. All of these isolates were confirmed as PcG sensitive with clover-leaf test. Internal controls showed little variation and external control isolates showed full agreement with the results obtained from a Danish study, suggesting that PcG zone diameter of ≥ 26 mm in combination with cloverleaf test can be used to detect penicillin susceptibility of S. aureus.    In conclusion, this study shows that nearly 1 /3 of the blood culture isolates of S. aureus from Kalmar are sensitive to benzylpenicillin.

Diss. (sammanfattning) Göteborg : Göteborgs universitet, 2007. / Härtill 3 uppsatser.

Thesis (M. D.)--University of New South Wales, 2003. / Also available online.

Thesis (M.S.)--University of Wisconsin--Madison, 1964. / eContent provider-neutral record in process. Description based on print version record. Bibliography: l. 90-105.

Thesis (Ph. D., Microbiology, Molecular Biology, and Biochemistry)--University of Idaho, July 2007. / Major professor: Gregory A. Bohach. Includes bibliographical references. Also available online (PDF file) by subscription or by purchasing the individual file.

Würzburg, Univ., Diss., 2009. / Zsfassung in engl. Sprache.

Thesis (Ph. D.)--Illinois State University, 1992. / Title from title page screen, viewed February 6, 2006. Dissertation Committee: Brian J. Wilkinson (chair), Radheshyam Jayaswal, Herman E. Brockman, Robert L. Preston, Philip D. Morse. Includes bibliographical references (leaves 107-112) and abstract. Also available in print.

Universiẗat, Diss., 2005--Würzburg.

Universiẗat, Diss., 2005--Würzburg.