Fifty three Staphylococcus aureus isolates were obtained from three centres, two hospitals and a private pathology laboratory, and studied for susceptibility to bacteriophages, resistance to antimicrobial agents and plasmid contents.Results of bacteriophage typing revealed that they belonged to a variety of phage types. Eighteen were untypable by any of the International Set of Phages, 16 belonged to phage group 111, nine to group I, four to group 11, two to group IV and two to the miscellaneous group.The isolates were resistant to one or more of methicillin (Mc), benzyl penicillin (Pc), gentamicin (Gm) , kanamycin (Km) , neomycin (Nm) , streptomycin (Sm) , trimethoprim (Tp), sulphonamides (Su), tetracycline (Tc), minocycline (Mn), chloramphenicol (Cm), novobiocin (Nb) and fusidic acid (Fa). Resistance to Pc was due to the production of beta-lactamase (Bla). No resistance to vancomycin, spectinomycin and erythromycin was detected. Resistance was also found to heavy metals such as cadmium (Cd), mercury (Hg), phenyl mercuric acetate (Pma), arsenate (Asa) and to the nucleic-acid binding compounds propamidine isethionate (Pi) and ethidium bromide (Eb).All but one of the isolates harboured plasmids. The number of plasmids the isolates carried varied from one to six and their sizes ranged from < 1.0 kb to c.48 kb.Location of the resistance determinants was ascertained by curing and transfer experiments. Loss of resistance was tested after growth at 43.5°C and transfer of resistance determinants was attempted by transduction, mixed-culture transfer and conjugation. The results revealed that resistance to Mc, Gm, Tp, Mn and Fa was chromosomal in all the resistant isolates and in some isolates Bla and resistance to Sm and Cd were chromosomal as well as plasmid encoded. In the majority of cases Bla and resistance to Km, Nm, Sm, Tc, Cin, Cd, Hg, Asa, Pma, Pi and Eb was encoded ++ / by plasmids.Conjugation experiments led to the isolation of three unique conjugative plasmids which have not been found to confer resistance to antimicrobials or to produce haemolysins or diffusible pigment (Dip). The three plasmids, pWBG620, pWBG637 and pWBG661, were indistinguishable by restriction endonuclease analysis and DNA-DNA hybridisation. However pWBG620, unlike pWBG637 and pWBG661, was not detected in the cytoplasm of its host and was only detected in transconjugants after it mobilised a non-conjugative Sm-resistance (SmR) plasmid. Further analysis indicated that it is integrated into the chromosome of its host, excises during conjugation and mobilises the SmR plasmid.These plasmids were studied further using pWBG637 as a representative. It was compared with representatives of the two groups of conjugative plasmids which have been reported in the staphylococci. These are the plasmids which encode resistance to Gm, Km and Nm and those which code for the production of diffusible pigment. The three types of conjugative plasmids were compared by restriction endonuclease analysis and DNA- DNA hybridisation and were found to be different. A preliminary restriction map of pWBG637 has also been constructed.However since pWBG637 has no resistance phenotype direct selection for it was not possible in transfer experiments and for incompatibility (Inc.). To study it further it was necessary to construct resistant derivatives which could be selected for in transfer experiments. This was achieved by labelling pWBG637 with resistance transposons to generate two conjugative plasmids, pWBG636 carrying an insert of Tn3851 (Gm- resistance) and pWBG642 carrying an insert of Tn551 (hn- resistance). It was found that transposon labelling had not changed the incompatibility of pWBG637 and therefore pWBG636 and pWBG642 were used in further experiments in place of pWBG637. Inc. ++ / tests with the pWBG637 derivatives revealed that the pWBG637 type of plasmid is not only different from the other two types of conjugative plasmids but is different from any of the described staphylococcal Inc. groups and therefore the pWBG637 type of plasmids represent a new Inc. group 15. The pWBG637 type of plasmids were studied further using plasmids pWBG636 and pWBG642. They were able to transfer conjugatively to a capsulated S.aureus strain either by the polyethylene glycol method or on filter membranes. They also transferred by conjugation to S. epidermidis and Streptococcus faecalis and were able to transfer back from these strains to S.aureus indicating that they also replicate in these hosts. Consequently they have been used to mobilise non-conjugative plasmids from S.epidermidis and non phage typable S.aureus. Both plasmids failed to transfer conjugatively to Bacillus subtilis and Escherichia coli.pWBG637 transferred non-conjugative plasmids by mobilising them in a manner similar to mobilisation (donation) in E.coli or by recombining with them to form new resistance plasmids. In one case, pWBG628 which encodes Bla and resistance to Cd, Km, Nm and Sm and has no homology with pWBG637 recombined with it during conjugation to produce three new conjugative plasmids pWBG629, pWBG630 and pWBG631 carrying resistance determinants from pWBG628. One of these plasmids, pWBG629, was found to be pWBG637 which had acquired a 4.5 kb element encoding resistance to Km, Nm and Sm. This element was shown to be transposable in both rec+ and rec- backgrounds and has been designated Tn3854. It expressed Sm resistance in E.coli and differs on this account from the Gram-negative transposon Tn5 which expresses resistance to Km, Nm and Sm in non-enteric bacteria but only resistance to Km and Nm in E. coli.Where possible the non-conjugative plasmids encoding resistance to ++ / antimicrobial agents were compared with phenotypically similar plasmids isolated from other parts of the world. It was found that the Tc and Sm resistance plasmids were closely related to other plasmids with the same phenotype whereas the Cm resistance plasmids were different.Although the majority of the Bla plasmids belonged to Inc. group 1 they demonstrated significant restriction enzyme fragment length polymorphism when compared with other Bla plasmids.This study has provided the first data on the genetics of antimicrobial resistance in Nigerian S.aureus. Although many of the plasmids studied were found to be similar to those previously described the isolates also contained some unique and previously undescribed plasmids.

The Saccharomyces cerevisiae plasmid, 2 micron circle, resides in the yeast nucleus at a high copy number. It provides no apparent growth advantage to its host, nor imposes any significant growth disadvantage. The plasmid is an excellent paradigm for studying mechanisms utilized in the persistence of a eukaryotic selfish DNA element that is selectively neutral. The plasmid achieves stable propagation and copy number maintenance by combining a partitioning system and an amplification system. The partitioning proteins Rep1p and Rep2p promote the recruitment of the histone H3 variant Cse4p and the yeast cohesin complex to the partitioning locus STB during S phase, leading to the formation of a functional partitioning complex which segregates the plasmid equally during mitosis. The integrity of the mitotic spindle is a pre-requisite for the specific nuclear localization of the plasmid as well as for plasmid association with a subset of the partitioning proteins such as Cse4p and the cohesin complex. The work presented in this thesis reveals, using tools of molecular genetics and cell biology, the involvement and possible functions of a microtubule associated nuclear motor protein, Kip1p, in the 2 micron circle partitioning pathway. The plasmid missegregates in kip1[Delta] cells, but not in cells harboring deletions of genes coding for the other nuclear motors. Kip1p interacts with the plasmid partitioning system and promotes the association of Cse4p and the cohesin complex with STB. Lack of Kip1p function delocalizes the plasmid from its characteristic nuclear locale in close proximity to the spindle pole body. The distance between a reporter plasmid and the spindle pole body is nearly doubled in a kip1[Delta] host strain. We propose that, unlike the conventional roles played by nuclear motors in spindle function and chromosome segregation, the Kip1p motor assists the 2 micron circle in associating with the mitotic spindle and translocating to its ‘partitioning center’. / text

No description available.

RNA b is the most abundant member of a family of autonomously replicating single- and double-stranded RNA plasmids found in mitochondria of several maize races. The extent to which this molecule associates in vivo with proteins was investigated by both rate zonal and CsCl equilibrium density gradient centrifugations of clarified lysate of mitochondria from maize plants with the S-type cytoplasm. A soluble endogenous complex of RNA b, responsible for synthesis of the more abundant (+) RNA b strand in in vitro conditions (in mitochondrial lysate), was identified. The complex had a density of 1.49 g/cm$ sp3$ but a surprisingly low sedimentation coefficient, only slightly larger than the naked RNA b. Only a minor fraction of RNA b molecules were bound in the complex; the majority of RNA b sedimented as naked RNA molecules. Complexes synthesizing other, less abundant, maize RNA plasmids were not identified. However, in vitro synthesis of all RNA plasmid species in mitochondrial lysate was resistant to heparin, suggesting that in all cases preformed RdRp-RNA template complexes, capable of elongating in vivo preinitiated RNA plasmid strands, were present.

S. aureus 1054. The recombination occurs by a novel method. The data show that pSl or pΔD contribute the site for recombination and that the gene(s) for the protein(s) involved in recombination are encoded on pOX1054 or the 1054 chromosome. Integration of the plasmids into the chromosome of 1054 was not detected.

Diss. (sammanfattning) Uppsala : Sveriges lantbruksuniv., 2002. / Härtill 4 uppsatser.

Thesis (Ph. D.)--University of Texas at Austin, 2008. / Vita. Includes bibliographical references.

Thesis (M.Sc.)--Memorial University of Newfoundland, 1997. / Restricted until May 1998. Bibliography: leaves 90-102.

Thesis (Ph. D.)--Carleton University, 1994. / Also available in electronic format on the Internet.

Thesis--Wisconsin. / Vita. Includes bibliographical references (leaves 296-301).